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100

21952165

What is the function of the AIRE gene at the embryonic stage?

Aire controls mesenchymal stem cell-mediated suppression in chronic colitis. Mesenchymal stem cells (MSCs) are emerging as a promising immunotherapeutic, based largely on their overt suppression of T lymphocytes under inflammatory and autoimmune conditions. While paracrine cross-talk between MSCs and T cells has been well-studied, an intrinsic transcriptional switch that programs MSCs for immunomodulation has remained undefined. Here we show that bone marrow-derived MSCs require the transcriptional regulator Aire to suppress T cell-mediated pathogenesis in a mouse model of chronic colitis. Surprisingly, Aire did not control MSC suppression of T cell proliferation in vitro. Instead, Aire reduced T cell mitochondrial reductase by negatively regulating a proinflammatory cytokine, early T cell activation factor (Eta)-1. Neutralization of Eta-1 enabled Aire(-/-) MSCs to ameliorate colitis, reducing the number of infiltrating effector T cells in the colon, and normalizing T cell reductase levels. We propose that Aire represents an early molecular switch imposing a suppressive MSC phenotype via regulation of Eta-1. Monitoring Aire expression in MSCs may thus be a critical parameter for clinical use.

101

22152485

What is the principle of the PAR-CLIP methodology?

In vivo and transcriptome-wide identification of RNA binding protein target sites. Animal mRNAs are regulated by hundreds of RNA binding proteins (RBPs). The identification of RBP targets is crucial for understanding their function. A recent method, PAR-CLIP, uses photoreactive nucleosides to crosslink RBPs to target RNAs in cells prior to immunoprecipitation. Here, we establish iPAR-CLIP (in vivo PAR-CLIP) to determine, at nucleotide resolution, transcriptome-wide binding sites of GLD-1, a conserved, germline-specific translational repressor in C. elegans. We identified 439 reproducible target mRNAs and demonstrate an excellent dynamic range of target detection by iPAR-CLIP. Upon GLD-1 knockdown, protein but not mRNA expression of the 439 targets was specifically upregulated, demonstrating functionality. Finally, we discovered strongly conserved GLD-1 binding sites near the start codon of target genes. These sites are functional in vitro and likely confer strong repression in vivo. We propose that GLD-1 interacts with the translation machinery near the start codon, a so-far-unknown mode of gene regulation in eukaryotes.

102

16910349

Which drugs are utilized to treat amiodarone-induced thyroitoxicosis?

Total thyroidectomy in amiodarone-induced thyrotoxicosis. Preoperative, intraoperative and postoperative considerations. A female patient was admitted to our Department for total thyroidectomy in amiodarone-induced thyrotoxicosis. The drug was prescribed for ventricular arrhythmia and atrial paroxysmal fibrillation in dilated cardiomyopathy due to chronic aortic regurgitation with left ventricular dysfunction (ejection fraction 35%; Class Functional NYHA III) and moderate-severe respiratory insufficiency. The cardiologist-anesthetist team has allowed to evaluate the surgical-cardiovascular-anesthesiologic risks and the balance between the improvement by the amiodarone administration for the arrhythmia, and the discontinuation of this treatment in order to prevent aggravation of the thyrotoxicosis. These hypotheses were subsequently discharged for the two reasons listed below: - several other antiarrhytmic drugs (that didn't show equivalent efficacy as amiodarone in preventing or converting such ventricular and atrial arrhythmias) may be proposed in the place of amiodarone. However, this could expose the patient to an arrhythmia; - a clear proof that the suspension of amiodarone can allow restoring normalization of the thyroid function doesn't exist. Therefore, the patient has been successfully submitted to the surgical intervention and in the follow-up we brought her back to a state of normalized thyroid function and cardiovascular conditions. In patients that cannot safely discontinue amiodarone or when medical therapy is ineffective in controlling thyrotoxicosis, thyroidectomy is the treatment of choice.

103

2816867

How is spastic diplegia diagnosed?

Early diagnosis of spastic diplegia, spastic hemiplegia, and quadriplegia. A retrospective study examined early neurodevelopmental behaviors of children with spastic diplegia, spastic hemiplegia, and quadriplegia (spastic, athetoid, or mixed) who had been followed up longitudinally in a high-risk infant follow-up clinic. Compared with peers with normal outcomes, children with all three types of cerebral palsy had significantly lower scores on the Bayley Mental Scale at 4 months of age; children with hemiplegia and quadriplegia also scored significantly lower on the Bayley Motor Scale. On the Movement Assessment of Infants at 4 months of age, the children with hemiplegia and quadriplegia showed significantly higher risk scores than the nonhandicapped group. The Movement Assessment of Infants was more than three times as sensitive as the Bayley Motor Scale in detecting motor abnormalities in 4-month-old infants with diplegia and more than twice as sensitive in detecting early abnormalities of hemiplegia. At 1 year of age, however, the Bayley Motor Scale was extremely sensitive in picking up motor deficits in children with all three types of cerebral palsy.

104

16835897

Which is the genetic defect causing Neurofibromatosis type 1?

Identification of forty-five novel and twenty-three known NF1 mutations in Chinese patients with neurofibromatosis type 1. Neurofibromatosis type 1 (NF1), characterized by skin neurofibromas and an excess of café-au-lait spots, is due to mutations in the neurofibromin (NF1) gene. Identifying the genetic defect in individuals with the disease represents a significant challenge because the gene is extremely large with a high incidence of sporadic mutations across the entire gene ranging from single nucleotide substitutes to large deletions. In the present study, we have used a combination of techniques (heteroduplex analysis, sequencing, loss of heterozygosity and quantification of gene dosage) to define the genetic defect in 68 individuals from a cohort of 107 NF1 Taiwanese patients of Chinese origin. Fifty-eight were initially identified using heteroduplex analytical techniques and confirmed by sequence analysis. A further five were identified by direct sequence analysis alone. The reminders were shown to carry large deletions in the NF1 gene by demonstrating loss of heterozygosity that was confirmed by gene dosage measurements using quantitative-PCR techniques. Mis-sense, non-sense, frame-shift or splice-site mutations were identified across the entire gene of which the majority (45/68) were novel in nature. The detection rate with the various analytical techniques and the types of mutation detected are consistent with published data involving both individuals and large cohort studies from other ethnic backgrounds.

105

10692426

Which is the human selenoprotein that contains several Se-Cys residues?

Selenoprotein P expression, purification, and immunochemical characterization. Most selenoproteins contain a single selenocysteine residue per polypeptide chain, encoded by an in-frame UGA codon. Selenoprotein P is unique in that its mRNA encodes 10-12 selenocysteine residues, depending on species. In addition to the high number of selenocysteines, the protein is cysteine- and histidine-rich. The function of selenoprotein P has remained elusive, in part due to the inability to express the recombinant protein. This has been attributed to presumed inefficient translation through the selenocysteine/stop codons. Herein, we report for the first time the expression of recombinant rat selenoprotein P in a transiently transfected human epithelial kidney cell line, as well as the endogenously expressed protein from HepG2 and Chinese hamster ovary cells. The majority of the expressed protein migrates with the predicted 57-kDa size of full-length glycosylated selenoprotein P. Based on the histidine-rich nature of selenoprotein P, we have purified the recombinant and endogenously expressed proteins using nickel-agarose affinity chromatography. We show that the recombinant rat and endogenous human proteins react in Western blotting and immunoprecipitation assays with commercial anti-histidine antibodies. The ability to express, purify, and immunochemically detect the recombinant protein provides a foundation for investigating the functions and efficiency of expression of this intriguing protein.

106

23671339

Which package is available for analysing genomic interactions in R/Bioconductor?

r3Cseq: an R/Bioconductor package for the discovery of long-range genomic interactions from chromosome conformation capture and next-generation sequencing data. The coupling of chromosome conformation capture (3C) with next-generation sequencing technologies enables the high-throughput detection of long-range genomic interactions, via the generation of ligation products between DNA sequences, which are closely juxtaposed in vivo. These interactions involve promoter regions, enhancers and other regulatory and structural elements of chromosomes and can reveal key details of the regulation of gene expression. 3C-seq is a variant of the method for the detection of interactions between one chosen genomic element (viewpoint) and the rest of the genome. We present r3Cseq, an R/Bioconductor package designed to perform 3C-seq data analysis in a number of different experimental designs. The package reads a common aligned read input format, provides data normalization, allows the visualization of candidate interaction regions and detects statistically significant chromatin interactions, thus greatly facilitating hypothesis generation and the interpretation of experimental results. We further demonstrate its use on a series of real-world applications.

107

20821198

How many clinical trials for off-label drugs in neonates are cited in the literature.

Paediatric investigation plans for pain: painfully slow! PURPOSE: To examine the early impact of the Paediatric Regulation, which entered into force in Europe on 27 January 2007, on the development of pharmaceutical drugs in the therapeutic field of pain submitted to the Paediatric Committee (PDCO) and to the European Medicines Agency (EMA). METHODS: Paediatric Investigations Plans (PIPs) submitted with a Decision (outcome) reached between September 2007 and March 2010 were included in the analysis. RESULTS: Of the 17 Paediatric Investigation Plans submitted, 14 have resulted in an EMA Decision, 3 were withdrawn by the applicants, 8 were granted a full waiver from development, and 1 resulted in a negative opinion. Decisions as issued included 15 clinical trials, with at least 1,282 children to be recruited into studies across five different products. Neonates were included in four of the products. CONCLUSIONS: The small number of submissions indicates a lack of new drugs being developed for the management of pain. Ethical concerns that too many vulnerable children will be recruited into clinical trials must be balanced against limiting the number of off-label prescribing and obtaining age-appropriate information on paediatric use. Now is an opportune time for clinicians, academics, learned societies and industry to collaborate for the benefit of children in pain.

108

23474818

Are stress granules involved in the pathogenesis of Amyotrophic Lateral Sclerosis?

ALS mutant FUS disrupts nuclear localization and sequesters wild-type FUS within cytoplasmic stress granules. Mutations in the gene encoding Fused in Sarcoma (FUS) cause amyotrophic lateral sclerosis (ALS), a fatal neurodegenerative disorder. FUS is a predominantly nuclear DNA- and RNA-binding protein that is involved in RNA processing. Large FUS-immunoreactive inclusions fill the perikaryon of surviving motor neurons of ALS patients carrying mutations at post-mortem. This sequestration of FUS is predicted to disrupt RNA processing and initiate neurodegeneration. Here, we demonstrate that C-terminal ALS mutations disrupt the nuclear localizing signal (NLS) of FUS resulting in cytoplasmic accumulation in transfected cells and patient fibroblasts. FUS mislocalization is rescued by the addition of the wild-type FUS NLS to mutant proteins. We also show that oxidative stress recruits mutant FUS to cytoplasmic stress granules where it is able to bind and sequester wild-type FUS. While FUS interacts with itself directly by protein-protein interaction, the recruitment of FUS to stress granules and interaction with PABP are RNA dependent. These findings support a two-hit hypothesis, whereby cytoplasmic mislocalization of FUS protein, followed by cellular stress, contributes to the formation of cytoplasmic aggregates that may sequester FUS, disrupt RNA processing and initiate motor neuron degeneration.

109

12374778

Does TGF-beta play a role in cardiac regeneration after myocardial infarction?

Stem cell differentiation requires a paracrine pathway in the heart. Members of the transforming growth factor beta1 (TGF-beta) superfamily--namely, TGF-beta and BMP2--applied to undifferentiated murine embryonic stem cells up-regulated mRNA of mesodermal (Brachyury) and cardiac specific transcription factors (Nkx2.5, MEF2C). Embryoid bodies generated from stem cells primed with these growth factors demonstrated an increased potential for cardiac differentiation with a significant increase in beating areas and enhanced myofibrillogenesis. In an environment of postmitotic cardiomyocytes, stem cells engineered to express a fluorescent protein under the control of a cardiac promoter differentiated into fluorescent ventricular myocytes beating in synchrony with host cells, a process significantly enhanced by TGF-beta or BMP2. In vitro, disruption of the TGF-beta/BMP signaling pathways by latency-associated peptide and/or noggin prevented differentiation of stem cells. In fact, only host cells that secrete a TGF-beta family member induced a cardiac phenotype in stem cells. In vivo, transplantation of stem cells into heart also resulted in cardiac differentiation provided that TGF-beta/BMP2 signaling was intact. In infarcted myocardium, grafted stem cells differentiated into functional cardiomyocytes integrated with surrounding tissue, improving contractile performance. Thus, embryonic stem cells are directed to differentiate into cardiomyocytes by signaling mediated through TGF-beta/BMP2, a cardiac paracrine pathway required for therapeutic benefit of stem cell transplantation in diseased heart.

110

26060713

Is there a genetic component for happiness?

Happiness & Health: The Biological Factors- Systematic Review Article. Happiness underlying factors are considerable from two dimensions: endogenic factors (biological, cognitive, personality and ethical sub-factors) and exogenic factors (behavioral, socialcultural, economical, geographical, life events and aesthetics sub-factors). Among all endogenic factors, biological sub-factors are the significant predictors of happiness. Existence of significant differences in temperament and happiness of infants is an indicator of biological influences. Therefore, this study aimed to consider biological factors that underlie happiness. At the first, all of the biological factors in relation with happiness were searched from following websites: PubMed, Wiley& Sons, Science direct (1990-2014). Then, the articles divided into five sub-groups (genetic, brain and neurotransmitters, endocrinology and hormones, physical health, morphology and physical attractiveness). Finally, a systematic review performed based on existing information. Results of studies on genetic factors indicated an average effectiveness of genetic about 35 -50 percent on happiness. In spite of difficulties in finding special genes, several genes distributed to emotion and mood. Neuroscience studies showed that some part of brain (e.g. amygdala, hipocamp and limbic system) and neurotransmitters (e.g. dopamine, serotonin, norepinefrine and endorphin) play a role in control of happiness. A few studies pointed to the role of cortisol and adrenaline (adrenal gland) and oxitocin (pituitary gland) in controlling happiness. Physical health and typology also concluded in most related studies to have a significant role in happiness. Therefore, according to previous research, it can be said that biological and health factors are critical in underlying happiness and its role in happiness is undeniable.

111

24271646

What enzyme is inhibied by Opicapone?

Effect of moderate liver impairment on the pharmacokinetics of opicapone. PURPOSE: Opicapone (OPC) is a novel catechol-O-methyltransferase (COMT) inhibitor to be used as adjunctive therapy in levodopa-treated patients with Parkinson's disease. The purpose of this study was to evaluate the effect of moderate liver impairment on the pharmacokinetics (PK) and pharmacodynamics (PD; effect on COMT activity) of OPC. METHODS: An open-label, parallel-group study in patients (n = 8) with moderate liver impairment (Child-Pugh category B, score of 7 to 9) and matched healthy subjects (n = 8, control) with normal liver function. All subjects received a single 50-mg oral dose of OPC, with plasma and urine concentrations of opicapone and its metabolites measured up to 72 h post-dose, including soluble COMT (S-COMT) activity. A one-way analysis of variance (ANOVA) was used to compare the main PK and PD parameters between groups. Point estimates (PE) of geometric mean ratios (GMR) and corresponding 90 % confidence intervals (90%CI) for the ratio hepatic/control subjects of each parameter were calculated and compared with the reference interval (80-125 %). RESULTS: Exposure to opicapone (AUC and Cmax) increased significantly in patients with moderate hepatic impairment (PE [90%CI]: AUC0-∞, 184 % [135-250 %]; Cmax, 189 % [144-249 %]). Although apparent total clearance (CL/F) of opicapone was decreased by ∼35 %, similar elimination half-life and unbound/bound fractions of opicapone were observed between the two groups. Both rate and extent of exposure to BIA 9-1103 were higher in the hepatically impaired group, but not statistically significant compared with the control group. Similar to the parent (opicapone), the observed increase in exposure to BIA 9-1106 was statistically significant in the hepatically impaired group over the control group. BIA 9-1106 was the only metabolite detected in urine and its urine PK parameters were in accordance with plasma data. Maximum S-COMT inhibition (Emax) occurred earlier for the hepatically impaired group with values of 100 % and 91.2 % for the hepatically impaired and control groups respectively. Both Emax and AUEC for the hepatically impaired group reached statistical significance over the control group. OPC was well tolerated in both hepatically impaired and control groups. CONCLUSION: The bioavailability of an orally administered single dose of 50 mg OPC was significantly higher in patients with moderate chronic hepatic impairment, perhaps by a reduced first-pass effect. As the tolerability profile of OPC was favourable under the conditions of this study and its exposure is completely purged from systemic circulation before the subsequent dose administration, no OPC dose adjustment is needed in patients with mild to moderate chronic hepatic impairment. However, as OPC is under clinical development for use as adjunctive therapy in levodopa-treated patients with Parkinson's disease, an adjustment of levodopa and/or OPC regimens in patients should be carefully considered based on a potentially enhanced levodopa dopaminergic response and the associated tolerability.

112

15789345

What kind of affinity purification would you use in order to isolate soluble lysosomal proteins?

The human brain mannose 6-phosphate glycoproteome: a complex mixture composed of multiple isoforms of many soluble lysosomal proteins. The lysosome is a membrane delimited cytoplasmic organelle that contains at least 50 hydrolytic enzymes and associated cofactors. The biomedical importance of these enzymes is highlighted by the many lysosomal storage disorders that are associated with mutations in genes encoding lysosomal proteins, and there is also evidence that lysosomal activities may be involved in more widespread human diseases. The aim of this study was to characterize the human brain lysosomal proteome with the goal of establishing a reference map to investigate human diseases of unknown etiology and to gain insights into the cellular function of the lysosome. Proteins containing mannose 6-phosphate (Man6-P), a carbohydrate modification used for targeting resident soluble lysosomal proteins to the lysosome, were affinity-purified using immobilized Man6-P receptor. Fractionation by two-dimensional electrophoresis resolved a complex mixture comprising approximately 800 spots. Constituent proteins in each spot were identified using a combination of matrix-assisted laser desorption/ionization-time of flight mass spectrometry (both peptide mass fingerprinting and tandem mass spectrometry) [corrected] on in-gel tryptic digests and N-terminal sequencing. In a complementary analysis, we also analyzed a tryptic digest of the unfractionated mixture by liquid chromatography MS/MS. In total, 61 different proteins were identified. Seven were likely contaminants associated with true Man6-P glycoproteins. Forty-one were known lysosomal proteins of which 11 have not previously been reported to contain Man6-P. An additional nine proteins were either uncharacterized or proteins not previously reported to have lysosomal function. We found that the human brain Man6-P-containing lysosomal proteome is highly complex and contains more proteins with a much greater number of individual isoforms than found in previous studies of Man6-P glycoproteomes.

113

19718047

Which are the genes thought to be regulated by EWS/FLI?

GSTM4 is a microsatellite-containing EWS/FLI target involved in Ewing's sarcoma oncogenesis and therapeutic resistance. Ewing's sarcoma is a malignant bone-associated tumor of children and young adults. Most cases of Ewing's sarcoma express the EWS/FLI fusion protein. EWS/FLI functions as an aberrant ETS-type transcription factor and serves as the master regulator of Ewing's sarcoma-transformed phenotype. We recently showed that EWS/FLI regulates one of its key targets, NR0B1, through a GGAA-microsatellite in its promoter. Whether other critical EWS/FLI targets are also regulated by GGAA-microsatellites was unknown. In this study, we combined transcriptional analysis, whole genome localization data, and RNA interference knockdown to identify glutathione S-transferase M4 (GSTM4) as a critical EWS/FLI target gene in Ewing's sarcoma. We found that EWS/FLI directly binds the GSTM4 promoter, and regulates GSTM4 expression through a GGAA-microsatellite in its promoter. Reduction of GSTM4 levels caused a loss of oncogenic transformation. Furthermore, reduction of GSTM4 resulted in an increased sensitivity of Ewing's sarcoma cells to chemotherapeutic agents, suggesting a role for this protein in drug resistance. Consistent with this hypothesis, patients with Ewing's sarcoma whose tumors had higher levels of GSTM4 expression had worse outcomes than those with lower expression levels. These data show that GSTM4 contributes to the cancerous behavior of Ewing's sarcoma and define a wider role for GGAA-microsatellites in EWS/FLI function than previously appreciated. These data also suggest a novel therapeutic resistance mechanism, in which the central oncogenic abnormality directly regulates a resistance gene.

114

15197606

Do archaeal genomes contain one or multiple origins of replication?

Identification of replication origins in the genome of the methanogenic archaeon, Methanocaldococcus jannaschii. Methanocaldococcus jannaschii has been notorious as an archaeon in which the replication origins are difficult to identify. Although extensive efforts have been exerted on this issue, the locations of replication origins still remain elusive 7 years after the publication of its complete genome sequence in 1996. Ambiguous results were obtained in identifying the replication origins of M. jannaschii based on all theoretical and experimental approaches. In the genome of M. jannaschii, we found that an ORF (MJ0774), annotated as a hypothetical protein, is a homologue of the Cdc6 protein. The position of the gene is at a global minimum of the x component of the Z curve, i.e., RY disparity curve, which has been used to identify replication origins in other Archaea. In addition, an intergenic region (694,540-695,226 bp) that is between the cdc6 gene and an adjacent ORF shows almost all the characteristics of known replication origins, i.e., it is highly rich in AT composition (80%) and contains multiple copies of repeat elements and AT stretches. Therefore, these lines of evidence strongly suggest that the identified region is a replication origin, which is designated as oriC1. The analysis of the y component of the Z curve, i.e., MK disparity curve, suggests the presence of another replication origin corresponding to one of the peaks in the MK disparity curve at around 1,388 kb of the genome.

115

16922728

Which pathological conditions are caused by mutations in the CYLD gene?

CYLD mutations underlie Brooke-Spiegler, familial cylindromatosis, and multiple familial trichoepithelioma syndromes. Brooke-Spiegler syndrome (BSS), familial cylindromatosis (FC), and multiple familial trichoepithelioma (MFT), originally described as distinct inherited disorders, are characterized by a variety of skin appendage neoplasms. Mutations in the CYLD gene are found in individuals with these syndromes. We describe a single family with affected members exhibiting either the FC or the MFT phenotypes associated with a mutation in the CYLD gene. These findings support the notion that BSS, FC, and MFT represent phenotypic variation of a single defect. Of interest, one of the affected individuals described in this report exhibits a severe phenotype illustrating the morbidity of the disorder.

116

19062530

Which is the genetic basis of Spinal Muscular Atrophy (SMA)?

[Molecular genetic basis of proximal spinal muscular atrophy and experience in its pharmaceutical treatment]. The review considers the original and published data on the molecular genetic basis of proximal spinal muscular atrophy (SMA), the most common monogenic neuromuscular disease. The structures of the SMN1 gene and SMN2 pseudogene, mutations distorting the SMN1 function, the structure and functions of the Smn neurotrophic protein, its role in biogenesis of small nuclear ribonucleoproteins (snRNPs), and the principles and prdblems of molecular diagnosis in SMA are described. Special consideration is given to the current approaches and prospects of gene and cell therapy of SMA, pharmacogenetic methods to correct the SMN2 function, and original results of long-term treatment of SMA patients with valproic acid drugs.

117

24062268

Which are the common symptoms of Cushing's syndrome?

Cushing syndrome. Cushing syndrome is the constellation of signs and symptoms caused by protracted exposure to glucocorticoids. The most common cause of Cushing syndrome in children and adolescents is exogenous administration of glucocorticoids. Presenting features commonly include weight gain, growth retardation, hirsutism, obesity, striae, acne and hypertension. Almost invariably, linear growth is severely diminished, a factor which may be useful in differentiating between childhood obesity and Cushing syndrome. Diagnostic approaches are based on distinguishing between adrenocorticotropic hormone (ACTH)-dependent and ACTH-independent etiologies, and consideration of the most likely diagnosis by age. Treatment modality is dependent upon etiology. After cure, important components of care include attention to linear growth, pubertal progression and body composition.

118

22795129

Which is the third subunit of the TSC1-TSC2 complex upstream of mTORC1?

TBC1D7 is a third subunit of the TSC1-TSC2 complex upstream of mTORC1. The tuberous sclerosis complex (TSC) tumor suppressors form the TSC1-TSC2 complex, which limits cell growth in response to poor growth conditions. Through its GTPase-activating protein (GAP) activity toward Rheb, this complex inhibits the mechanistic target of rapamycin (mTOR) complex 1 (mTORC1), a key promoter of cell growth. Here, we identify and biochemically characterize TBC1D7 as a stably associated and ubiquitous third core subunit of the TSC1-TSC2 complex. We demonstrate that the TSC1-TSC2-TBC1D7 (TSC-TBC) complex is the functional complex that senses specific cellular growth conditions and possesses Rheb-GAP activity. Sequencing analyses of samples from TSC patients suggest that TBC1D7 is unlikely to represent TSC3. TBC1D7 knockdown decreases the association of TSC1 and TSC2 leading to decreased Rheb-GAP activity, without effects on the localization of TSC2 to the lysosome. Like the other TSC-TBC components, TBC1D7 knockdown results in increased mTORC1 signaling, delayed induction of autophagy, and enhanced cell growth under poor growth conditions.

119

17457979

Which kinase is inhibited by the small molecule KN-93?

KN-93, a specific inhibitor of CaMKII inhibits human hepatic stellate cell proliferation in vitro. AIM: To investigate the effects of KN-93, a CaMKII selective inhibitor on cell proliferation and the expression of p53 or p21 protein in human hepatic stellate cells. METHODS: Human hepatic stellate cells (LX-2) were incubated with various concentrations (0-50 micromol/L) of KN-93 or its inactive derivative, KN-92. Cell proliferation was measured by CCK-8 assay, and the expression of two cell cycle regulators, p53 and p21, was determined by SDS-PAGE and Western blotting. RESULTS: KN-93 (5-50 micromol/L) decreased the proliferation of human hepatic stellate cells in a dose-dependent manner from 81.76% (81.76% +/- 2.58% vs 96.63% +/- 2.69%, P < 0.05) to 27.15% (27.15% +/- 2.86% vs 96.59% +/- 2.44%, P < 0.01) after 24 h treatment. Incubation of 10 micromol/L KN-93 induced the cell growth reduction in a time-dependent manner from 78.27% at 8 h to 11.48% at 48 h. However, KN-92, an inactive derivative of KN-93, did not inhibit cell proliferation effectively. Moreover, analysis of cell cycle regulator expression revealed that KN-93 rather than KN-92 reduced the expression of p53 and p21. CONCLUSION: KN-93 has potent inhibitory effect on proliferation of LX-2 cells by modulating the expression of two special cell cycle regulators, p53 and p21.

120

20080130

What is the effect of Chk2 splice variants on wild-type Chk2 kinase activity?

Chk2 splice variants express a dominant-negative effect on the wild-type Chk2 kinase activity. While the majority of RNA transcripts from protein-encoding genes in the human genome are subject to physiological splicing, pathological splicing is increasingly reported in cancer tissue. Previously, we identified >90 different splice variants of Chk2, a gene encoding a serine/threonine kinase propagating the DNA damage signal by phosphorylating and activating several downstream substrates like p53, Cdc25A, and Cdc25C involved in cell cycle arrest and apoptosis. While alternative splice forms of other genes have been reported to exert a dominant-negative effect on the wild-type molecules, the function of Chk2 splice protein variants is still unclear. Here we evaluated the function of four Chk2 splice proteins for which mRNA splice variants were identified in human breast carcinomas. These splice variants were stably expressed as nuclear proteins. Two splice forms (Chk2Delta4 and Chk2del(2-3)) expressed kinase activity while variants Chk2Delta11 and Chk2isoI were essentially kinase inactive. Independent of intrinsic kinase activity, each splice variant impaired wild-type Chk2 activity through heterodimerization. Based on our findings, we suggest alternative splicing as a possible novel mechanism for repression of the Chk2 wild-type function.

121

25522693

List genes that have been found mutated in CMT1A (Charcot-Marie-Tooth disease type 1 A).

PMP22-Related neuropathies and other clinical manifestations in Chinese han patients with charcot-marie-tooth disease type 1. INTRODUCTION: Most cases of Charcot-Marie-Tooth (CMT) disease are caused by mutations in the peripheral myelin protein 22 gene (PMP22), including heterozygous duplications (CMT1A), deletions (HNPP), and point mutations (CMT1E). METHODS: Single-nucleotide polymorphism (SNP) arrays were used to study PMP22 mutations based on the results of multiplex ligation-dependent probe amplification (MLPA) and polymerase chain reaction-restriction fragment length polymorphism methods in 77 Chinese Han families with CMT1. PMP22 sequencing was performed in MLPA-negative probands. Clinical characteristics were collected for all CMT1A/HNPP probands and their family members. RESULTS: Twenty-one of 77 CMT1 probands (27.3%) carried duplication/deletion (dup/del) copynumber variants. No point mutations were detected. SNP array and MLPA seem to have similar sensitivity. Fifty-seven patients from 19 CMT1A families had the classical CMT phenotype, except for 1 with concomitant CIDP. Two HNPP probands presented with acute ulnar nerve palsy or recurrent sural nerve palsy, respectively. CONCLUSIONS: The SNP array has wide coverage, high sensitivity, and high resolution and can be used as a screening tool to detect PMP22 dup/del as shown in this Chinese Han population.

122

18039618

Which viruses are best known to cause myocarditis?

[Molecular biological virus identification in dilated cardiomyopathy]. UNLABELLED: Enteroviruses have been considered to be the most common cause of acute myocarditis and possible consequence of dilated cardiomyopathy. Some publications shed light to the role of other viruses in this disease as well. Our molecular investigation has demonstrated that adeno- and herpes viruses might also frequently occur in dilated cardiomyopathy. AIM: The aim of our study was to screen virus genomes in heart tissues from heart-transplanted patients to prove their possible role in the pathogenesis of dilated cardiomyopathy. METHODS: DNA and RNA were isolated from five regions of the heart muscle. Amplification for Adenovirus Type 3, Human Herpes Virus Type 6 and Enterovirus genomes were performed by nested-Polymerase Chain Reaction. Finally the virus-positive samples were direct sequenced. RESULTS: In 2 patients Adenovirus Type 3 and in 1 patient both Adenovirus Type 3 and Human Herpes Virus Type 6 were detected. No enteroviruses were found in any heart tissue. CONCLUSIONS: In our study the adenovirus genome was found to be the most frequent virus genome in explanted heart tissues. The identified viral sequences proved previous viral infection, which could have played a role in the development of dilated cardiomyopathy. Detection of different viruses in the myocardium by molecular biological examinations might contribute to adequate treatment of these patients.

123

25122144

Which genes have been associated with Cerebral Cavernous Malformation?

Exceptional aggressiveness of cerebral cavernous malformation disease associated with PDCD10 mutations. PURPOSE: The phenotypic manifestations of cerebral cavernous malformation disease caused by rare PDCD10 mutations have not been systematically examined, and a mechanistic link to Rho kinase-mediated hyperpermeability, a potential therapeutic target, has not been established. METHODS: We analyzed PDCD10 small interfering RNA-treated endothelial cells for stress fibers, Rho kinase activity, and permeability. Rho kinase activity was assessed in cerebral cavernous malformation lesions. Brain permeability and cerebral cavernous malformation lesion burden were quantified, and clinical manifestations were assessed in prospectively enrolled subjects with PDCD10 mutations. RESULTS: We determined that PDCD10 protein suppresses endothelial stress fibers, Rho kinase activity, and permeability in vitro. Pdcd10 heterozygous mice have greater lesion burden than other Ccm genotypes. We demonstrated robust Rho kinase activity in murine and human cerebral cavernous malformation vasculature and increased brain vascular permeability in humans with PDCD10 mutation. Clinical phenotype is exceptionally aggressive compared with the more common KRIT1 and CCM2 familial and sporadic cerebral cavernous malformation, with greater lesion burden and more frequent hemorrhages earlier in life. We first report other phenotypic features, including scoliosis, cognitive disability, and skin lesions, unrelated to lesion burden or bleeding. CONCLUSION: These findings define a unique cerebral cavernous malformation disease with exceptional aggressiveness, and they inform preclinical therapeutic testing, clinical counseling, and the design of trials.Genet Med 17 3, 188-196.

124

18954857

Is DITPA a thyroid hormone analog utilized in experimental and clinical studies

[Thyroid hormone analogs: an important biological supply and new therapeutic possibilities]. Thyroid hormones [predominantly 3, 5, 3 -I- iodothyronine (T3)] regulate cholesterol and lipoprotein metabolism but cardiac effects restrict their use as hypolipidemic drugs. New molecules have been developped which target specifically the thyroid hormone receptor ss, predominant isoform in liver. The first thyroid hormone agonist, called GC1, has selective actions compared to T3. In animals, GC1 reduced serum cholesterol and serum triglycerides, probably by stimulation important steps in reverse cholesterol transport. Other selective thyromimetic, KB- 2115 and KB - 141 have similar effects. Another class of thyroid hormone analogs, the thyronamines have emerged recently but the basic biology of this new class of endogenous thyroid hormone remains to better understood. Therefore, these molecules may be a potentially treatment for obesity and reduction cholesterol, triglycerides and lipoprotein (a). To date the studies in human are preliminary. Tolerance and efficacy of these drugs are still under investigation.

125

20712856

What is Tarlov Cyst?

Confusion after spine injury: cerebral fat embolism after traumatic rupture of a Tarlov cyst: case report. BACKGROUND: Acute low back pain is a very common symptom and reason for many medical consultations. In some unusual circumstances it could be linked to a rare aetiology. CASE PRESENTATION: We report a 70-year-old man with an 8-month history of left posterior thigh and leg pain who had sudden confusion after a fall from standing. It was due to cerebral fat embolism suspected by computed tomography scan, later confirmed by brain magnetic resonance imaging (MRI). A spinal MRI scan was then performed and revealed a sacral fracture which drained into an unknown perineurial cyst (Tarlov cyst). Under medical observation the patient fully recovered within three weeks. CONCLUSIONS: Sacral perineurial cysts are rare, however they remain a potential cause of lumbosacral radiculopathy.

126

21431099

What are 'vildagliptin', 'sitagliptin', 'saxagliptin', 'alogliptin', 'linagliptin', and 'dutogliptin'?

Alogliptin for the treatment of type 2 diabetes. The pharmacologic management of type 2 diabetes has changed dramatically in the past two decades. We have moved from a situation of only having two choices, insulin and sulfonylureas, to a position of myriad choices from 11 categories of medications (insulin, sulfonylureas, biguanides, α-glucosidase inhibitors, gliptins (dipeptidyl peptidase 4 [DPP IV] inhibitors), bromocriptine, glucagon-like peptide analogues, thiazolidinediones, glinides, amylin analogues and bile acid sequestrants. One of the most recent additions to this list are the DPP IV inhibitors commonly known as gliptins. Currently, there are four DPP IV inhibitors available in various countries-alogliptin, sitagliptin, vildagliptin and saxagliptin (1). Of these, two have been approved for clinical use in the United States: sitagliptin and saxagliptin. Additionally, linagliptin, vildagliptin and alogliptin are currently in phase III development in the United States while studies with another DPP IV inhibitor, dutogliptin, have been terminated (2). Alogliptin was approved for use in Japan under the trade name Nesina® in April 2010 (3). This manuscript will review alogliptin, its chemistry, pharmacokinetics/pharmacodynamics, drug interactions, clinical trials and its current state of FDA review. Preclinical animal data have been reviewed elsewhere and will not be outlined in this manuscript. The interested reader is referred to those recent reviews (4, 5).

127

22560084

Which is the most important prognosis sub-classification in Chronic Lymphocytic Leukemia?

Mutational status and gene repertoire of IGHV-IGHD-IGHJ rearrangements in Serbian patients with chronic lymphocytic leukemia. UNLABELLED: The mutational status and configuration of immunoglobulin heavy variable (IGHV) gene rearrangements was analyzed in 85 Serbian patients with chronic lymphocytic leukemia (CLL). We found that 55.3% of cases belonged to mutated and 44.7% to unmutated CLL, progressive disease predominating in the unmutated subset. IGHV gene use resembled that obtained for Mediterranean countries, except for underrepresentation of the IGHV4 subgroup in our cohort. BACKGROUND: Chronic lymphocytic leukemia (CLL) results from the clonal expansion of mature B lymphocytes and is characterized by extreme clinical heterogeneity. One of the most reliable prognostic markers in chronic lymphocytic leukemia (CLL) is the mutational status of immunoglobulin heavy variable (IGHV) genes, which defines 2 subsets, mutated CLL (M-CLL) and unmutated CLL (U-CLL), with different clinical courses. Biased IGHV gene use between M-CLL and U-CLL clones, as well as population differences in the IGHV gene repertoire have been reported. PATIENTS AND METHODS: In this study, mutational status and configuration of IGHV-IGHD-IGHJ rearrangements in 85 Serbian patients were analyzed using reverse transcriptase-polymerase chain reaction (RT-PCR) and sequencing methodology. RESULTS: We found that 55.3% of cases belonged to M-CLL and 44.7% belonged to U-CLL, with progressive disease predominating in the unmutated subset. Most frequently expressed was the IGHV3 subgroup (55.7%), followed by IGHV1 (27.3%), IGHV4 (12.5%), IGHV5 (2.3%), IGHV2 (1.1%), and IGHV6 (1.1%). The distribution of IGHD subgroups was as follows: IGHD3, 39.1%; IGHD2, 21.8%; IGHD6, 12.6%; IGHD1, 10.3%; IGHD4, 8%; IGHD5, 6.9%; and IGHD7, 1.1%. The most frequent IGHJ gene was IGHJ4 (48.9%), followed by IGHJ6 (28.4%), IGHJ3 (11.4%), and IGHJ5 (11.4%). In 15.3% of cases, heavy complementarity-determining region 3 (VH CDR3) amino acid sequences could be assigned to previously defined stereotyped clusters. CONCLUSIONS: Our study showed a strong correlation between IGHV gene mutational status and clinical course of CLL. IGHV gene use was comparable to that obtained for Mediterranean countries, with the exception of the IGHV4 subgroup, which was underrepresented in our cohort.

128

19506735

Is MammaPrint cleared by the United States Food and Drug Administration?

Clinical utility of microarrays: current status, existing challenges and future outlook. Microarray-based clinical tests have become powerful tools in the diagnosis and treatment of diseases. In contrast to traditional DNA-based tests that largely focus on single genes associated with rare conditions, microarray-based tests are ideal for the study of diseases with underlying complex genetic causes. Several microarray based tests have been translated into clinical practice such as MammaPrint and AmpliChip CYP450. Additional cancer-related microarray-based tests are either in the process of FDA review or under active development, including Tissue of Tumor Origin and AmpliChip p53. All diagnostic microarray testing is ordered by physicians and tested by a Clinical Laboratories Improvement Amendment-certified (CLIA) reference laboratory. Recently, companies offering consumer based microarray testing have emerged. Individuals can order tests online and service providers deliver the results directly to the clients via a password-protected secure website. Navigenics, 23andMe and deCODE Genetics represent pioneering companies in this field. Although the progress of these microarray-based tests is extremely encouraging with the potential to revolutionize the recognition and treatment of common diseases, these tests are still in their infancy and face technical, clinical and marketing challenges. In this article, we review microarray-based tests which are currently approved or under review by the FDA, as well as the consumer-based testing. We also provide a summary of the challenges and strategic solutions in the development and clinical use of the microarray-based tests. Finally, we present a brief outlook for the future of microarray-based clinical applications.

129

15759228

Is amantadine effective for treatment of disorders conciousness?

Predictors of outcome in prolonged posttraumatic disorders of consciousness and assessment of medication effects: A multicenter study. OBJECTIVES: To develop predictive models of recovery from the vegetative state (VS) and minimally conscious state (MCS) after traumatic brain injury (TBI) and to gather preliminary evidence on the impact of various psychotropic medications on the recovery process to support future randomized controlled trials. Design Longitudinal observational cohort design, in which demographic information, injury and acute care history, neuroimaging data, and an initial Disability Rating Scale (DRS) score were collected at the time of study enrollment. Weekly follow-up data, consisting of DRS score, current psychoactive medications, and medical complications, were gathered until discharge from inpatient rehabilitation. SETTING: Seven acute inpatient rehabilitation facilities in the United States and Europe with specialized programs for treating patients in the VS and MCS. PARTICIPANTS: People with TBI (N=124) who were in the VS or MCS 4 to 16 weeks after injury. INTERVENTIONS: Not applicable. MAIN OUTCOME MEASURES: DRS score at 16 weeks after injury and time until commands were first followed (among those participants demonstrating no command following at study enrollment). Results DRS score at enrollment, time between injury and enrollment, and rate of DRS change during the first 2 weeks of poststudy observation were all highly predictive of both outcomes. No variables related to injury characteristics or lesions on neuroimaging were significant predictors. Of the psychoactive medications, amantadine hydrochloride was associated with greater recovery and dantrolene sodium was associated with less recovery, in terms of the DRS score at 16 weeks but not the time until commands were followed. More detailed analysis of the timing of functional improvement, with respect to the initiation of amantadine provided suggestive, but not definitive, evidence of the drug's causal role. CONCLUSIONS: These findings show the feasibility of improving outcome prediction from the VS and MCS using readily available clinical variables and provide suggestive evidence for the effects of amantadine and dantrolene, but these results require confirmation through randomized controlled trials.

130

25360794

What is needed for MMP proteins to be functional?

Circular trimers of gelatinase B/matrix metalloproteinase-9 constitute a distinct population of functional enzyme molecules differentially regulated by tissue inhibitor of metalloproteinases-1. Gelatinase B/matrix metalloproteinase-9 (MMP-9) (EC 3.4.24.35) cleaves many substrates and is produced by most cell types as a zymogen, proMMP-9, in complex with the tissue inhibitor of metalloproteinases-1 (TIMP-1). Natural proMMP-9 occurs as monomers, homomultimers and heterocomplexes, but our knowledge about the overall structure of proMMP-9 monomers and multimers is limited. We investigated biochemical, biophysical and functional characteristics of zymogen and activated forms of MMP-9 monomers and multimers. In contrast with a conventional notion of a dimeric nature of MMP-9 homomultimers, we demonstrate that these are reduction-sensitive trimers. Based on the information from electrophoresis, AFM and TEM, we generated a 3D structure model of the proMMP-9 trimer. Remarkably, the proMMP-9 trimers possessed a 50-fold higher affinity for TIMP-1 than the monomers. In vivo, this finding was reflected in a higher extent of TIMP-1 inhibition of angiogenesis induced by trimers compared with monomers. Our results show that proMMP-9 trimers constitute a novel structural and functional entity that is differentially regulated by TIMP-1.

131

24302690

What is hyperosmia

Pregnancy does not affect human olfactory detection thresholds. Hyperosmia is suspected in pregnancy; however, no empirical study using validated measures of olfactory function has clearly confirmed the anecdotal reports of this phenomenon. The goal of the current study is to compare the olfactory sensitivity of pregnant women to that of nonpregnant women and men. All participants rated their sense of smell and pregnant women listed the odors to which they were most sensitive. Detection thresholds were measured using a well-validated protocol. A group of pregnant and nonpregnant women was studied longitudinally using a signal detection procedure designed to detect small differences in sensitivity. Pregnant women, particularly in the 1st trimester, rated their sense of smell to be higher than nonpregnant women and men and indicated many (primarily unpleasant) odors to which they were more sensitive. Women rated their sense of smell higher than men. However, there was no sex difference in thresholds and neither thresholds nor signal detection measures of sensitivity were significantly affected by either sex or pregnancy status. The implications of the lack of relationship between self-report and measures of olfactory sensitivity, particularly in pregnancy, are discussed.

132

23126680

What is the number of long non coding RNAs in the human genome

Strategies to identify long noncoding RNAs involved in gene regulation. Long noncoding RNAs (lncRNAs) have been detected in nearly every cell type and found to be fundamentally involved in many biological processes. The characterization of lncRNAs has immense potential to advance our comprehensive understanding of cellular processes and gene regulation, along with implications for the treatment of human disease. The recent ENCODE (Encyclopedia of DNA Elements) study reported 9,640 lncRNA loci in the human genome, which corresponds to around half the number of protein-coding genes. Because of this sheer number and their functional diversity, it is crucial to identify a pool of potentially relevant lncRNAs early on in a given study. In this review, we evaluate the methods for isolating lncRNAs by immunoprecipitation and review the advantages, disadvantages, and applications of three widely used approaches - microarray, tiling array, and RNA-seq - for identifying lncRNAs involved in gene regulation. We also look at ways in which data from publicly available databases such as ENCODE can support the study of lncRNAs.

133

24961027

Which is the most known bacterium responsible for botulism (sausage-poisoning)?

Botulism as a food poisoning: what is it? Botulism is a rare but potentially life-threatening neuroparalytic syndrome resulting from the action of a neurotoxin elaborated by the microorganism Clostridium botulinum. This disease has a lengthy history; the first investigation of botulism occurred in the 1820s with a case report on hundreds of patients with "sausage poisoning" in a southern German town. Several decades later in Belgium, the association was demonstrated between a neuromuscular paralysis and ham infected by a spore forming bacillus that was isolated from the ham. The organism was named Bacillus botulinus after the Latin word for sausage, botulus.

134

25483063

What is the association of spermidine with α-synuclein neurotoxicity?

Spermidine protects against α-synuclein neurotoxicity. As our society ages, neurodegenerative disorders like Parkinson`s disease (PD) are increasing in pandemic proportions. While mechanistic understanding of PD is advancing, a treatment with well tolerable drugs is still elusive. Here, we show that administration of the naturally occurring polyamine spermidine, which declines continuously during aging in various species, alleviates a series of PD-related degenerative processes in the fruit fly Drosophila melanogaster and the nematode Caenorhabditis elegans, two established model systems for PD pathology. In the fruit fly, simple feeding with spermidine inhibited loss of climbing activity and early organismal death upon heterologous expression of human α-synuclein, which is thought to be the principal toxic trigger of PD. In this line, administration of spermidine rescued α-synuclein-induced loss of dopaminergic neurons, a hallmark of PD, in nematodes. Alleviation of PD-related neurodegeneration by spermidine was accompanied by induction of autophagy, suggesting that this cytoprotective process may be responsible for the beneficial effects of spermidine administration.

135

24190003

List symptoms of 4H leukodystrophy.

Teaching neuroimages: hypomyelinating leukodystrophy with hypodontia due to POLR3B: look into a leukodystrophy's mouth. An 18-year-old German woman presented with progressive cerebellar ataxia since early childhood, delayed cognitive development, and hypogonadotropic hypogonadism. MRI demonstrated diffuse cerebral hypomyelination, cerebellar atrophy, and thin corpus callosum; X-ray revealed persistent milk teeth and hypoplastic crowns and roots (figure), indicative of 4H syndrome (hypomyelination, hypodontia, hypogonadotropic hypogonadism). POLR3B sequencing(1) revealed 2 compound heterozygous mutations (C527R [C.1579T>C] and the common ancestral V523E [C.1568T>A](2)).

136

23539364

What is the extracellular core "matrisome"?

Gene evolution and functions of extracellular matrix proteins in teeth. The extracellular matrix (ECM) not only provides physical support for tissues, but it is also critical for tissue development, homeostasis and disease. Over 300 ECM molecules have been defined as comprising the "core matrisome" in mammals through the analysis of whole genome sequences. During tooth development, the structure and functions of the ECM dynamically change. In the early stages, basement membranes (BMs) separate two cell layers of the dental epithelium and the mesenchyme. Later in the differentiation stages, the BM layer is replaced with the enamel matrix and the dentin matrix, which are secreted by ameloblasts and odontoblasts, respectively. The enamel matrix genes and the dentin matrix genes are each clustered in two closed regions located on human chromosome 4 (mouse chromosome 5), except for the gene coded for amelogenin, the major enamel matrix protein, which is located on the sex chromosomes. These genes for enamel and dentin matrix proteins are derived from a common ancestral gene, but as a result of evolution, they diverged in terms of their specific functions. These matrix proteins play important roles in cell adhesion, polarity, and differentiation and mineralization of enamel and dentin matrices. Mutations of these genes cause diseases such as odontogenesis imperfect (OI) and amelogenesis imperfect (AI). In this review, we discuss the recently defined terms matrisome and matrixome for ECMs, as well as focus on genes and functions of enamel and dentin matrix proteins.

137

15579691

Is GAGA associated with nucleosome-free regions (NFR)?

The enhancer-blocking activity of the Fab-7 boundary from the Drosophila bithorax complex requires GAGA-factor-binding sites. In the work reported here we have analyzed the role of the GAGA factor [encoded by the Trithorax-like (Trl) gene] in the enhancer-blocking activity of Frontabdominal-7 (Fab-7), a domain boundary element from the Drosophila melanogaster bithorax complex (BX-C). One of the three nuclease hypersensitive sites in the Fab-7 boundary, HS1, contains multiple consensus-binding sequences for the GAGA factor, a protein known to be involved in the formation and/or maintenance of nucleosome-free regions of chromatin. GAGA protein has been shown to localize to the Fab-7 boundary in vivo, and we show that it recognizes sequences from HS1 in vitro. Using two different transgene assays we demonstrate that GAGA-factor-binding sites are necessary but not sufficient for full Fab-7 enhancer-blocking activity. We show that distinct GAGA sites are required for different enhancer-blocking activities at different stages of development. We also show that the enhancer-blocking activity of the endogenous Fab-7 boundary is sensitive to mutations in the gene encoding the GAGA factor Trithorax-like.

138

11353082

Which are the plant DNA (cytosine-5) methyltransferase families?

Two Arabidopsis methylation-deficiency mutations confer only partial effects on a methylated endogenous gene family. In Arabidopsis a SWI2/SNF2 chromatin remodeling factor-related protein DDM1 and a cytosine methyltransferase MET1 are required for maintenance of genomic cytosine methylation. Mutations in either gene cause global demethylation. In this work we have assessed the effects of these mutations on the PAI tryptophan biosynthetic gene family, which consists of four densely methylated genes arranged as a tail-to-tail inverted repeat plus two unlinked singlet genes. The methylation mutations caused only partial demethylation of the PAI loci: ddm1 had a strong effect on the singlet genes but a weaker effect on the inverted repeat, whereas met1 had a stronger effect on the inverted repeat than on the singlet genes. The double ddm1 met1 mutant also displayed partial demethylation of the PAI genes, with a pattern similar to the ddm1 single mutant. To determine the relationship between partial methylation and expression for the singlet PAI2 gene we constructed a novel reporter strain of Arabidopsis in which PAI2 silencing could be monitored by a blue fluorescent plant phenotype diagnostic of tryptophan pathway defects. This reporter strain revealed that intermediate levels of methylation correlate with intermediate suppression of the fluorescent phenotype.

139

18314594

Where is the histone variant CENPA preferentially localized?

[Embryonal germ cells and germ cell tumors]. Testicular germ cell tumors comprise of group of pluripotent tumors including seminomas and nonseminomas, arise from intratubular germ cell neoplasia and originate from the primordial germ cells/ gonocytes. Many well characterized markers of embryonic stem cells including CD9, PODXL and centromere-specific histone-H3-like protein CENPA are consistently expressed in TGCTs. In embryonic stem cells, pluripotency and self renewal capacities are provided by a network of OCT3/4, NANOG and SOX2. In testicular germ cell tumors, pluripotency genes OCT3/4 und NANOG are upregulated both, in seminomas and non-seminomas, while SOX2 is differentially upregulated in embryonal carcinomas only. Similar to embryonic stem cells, most histological elements of type II GCTs are sensitive to chemotherapy and irradiation. Furthermore, all invasive TGCTs show a consistent gain of the short arm of chromosome 12, as found in ES cells upon extensive in vitro culturing. Moreover, the genetic constitution of testicular germ cell tumors can also be linked to characteristics of embryonic stem cells, likely related to their specific inability to repair DNA damage and their high sensitivity to apoptotic cell death. In conclusion, testicular germ cell tumors represent embryonic cancers found in adults. Both the seminomas and nonseminomas have their specific population of stem cells representative of the primordial germ cells/gonocytes and for embryonic stem cells, respectively.

140

22705977

In which proteins is the chromodomain present?

Delayed fusion and altered gene expression contribute to semicircular canal defects in Chd7 deficient mice. Proper morphogenesis of inner ear semicircular canals requires precise regulation of cellular proliferation, epithelial-to-mesenchymal transition, and fusion of epithelial plates. Epigenetic regulation of these processes is not well understood, but is likely to involve chromatin remodeling enzymes. CHD7 is a chromodomain-containing, ATP dependent helicase protein that is highly expressed in the developing ear and is required for semicircular canal development in both humans and mice. Here we report that mice with heterozygous loss of Chd7 function exhibit delayed semicircular canal genesis, delayed Netrin1 expression and disrupted expression of genes that are critical for semicircular canal formation (Bmp2, Bmp4, Msx1 and Fgf10). Complete loss of Chd7 results in aplasia of the semicircular canals and sensory vestibular organs, with reduced or absent expression of Otx1, Hmx3, Jagged1, Lmo4, Msx1 and Sox2. Our results suggest that Chd7 may have critical selector gene functions during inner ear morphogenesis. Detailed analysis of the epigenetic modifications underlying these gene expression changes should provide insights into semicircular canal development and help in the design of therapies for individuals with inner ear malformations.

141

20185404

What is Genomicus?

Genomicus: a database and a browser to study gene synteny in modern and ancestral genomes. UNLABELLED: Comparative genomics remains a pivotal strategy to study the evolution of gene organization, and this primacy is reinforced by the growing number of full genome sequences available in public repositories. Despite this growth, bioinformatic tools available to visualize and compare genomes and to infer evolutionary events remain restricted to two or three genomes at a time, thus limiting the breadth and the nature of the question that can be investigated. Here we present Genomicus, a new synteny browser that can represent and compare unlimited numbers of genomes in a broad phylogenetic view. In addition, Genomicus includes reconstructed ancestral gene organization, thus greatly facilitating the interpretation of the data. AVAILABILITY: Genomicus is freely available for online use at http://www.dyogen.ens.fr/genomicus while data can be downloaded at ftp://ftp.biologie.ens.fr/pub/dyogen/genomicus.

142

15989900

Is amiodarone a class I anti-arrhythmic drug?

Amiodarone and the thyroid. Among the drugs affecting the thyroid gland, no drug has puzzled, and at the same time fascinated, endocrinologists more than amiodarone. Amiodarone is a potent class III anti-arrhythmic drug that also possesses beta-blocking properties. It is very rich in iodine, with a 100-mg tablet containing an amount of iodine that is 250 times the recommended daily iodine requirement. Amiodarone produces characteristic alterations in thyroid function tests in euthyroid patients. Understanding these alterations is crucial in avoiding unnecessary investigations and treatment. Amiodarone-induced thyroid dysfunction occurs because of both its iodine content and the direct toxic effects of the compound on thyroid parenchyma. Amiodarone-induced hyperthyroidism is more common in iodine-deficient regions of the world, whereas amiodarone-induced hypothyroidism is usually seen in iodine-sufficient areas. In contrast to amiodarone-induced hypothyroidism, amiodarone-induced thyrotoxicosis is a difficult condition to diagnose and treat. In this review, we discuss the alterations in thyroid function tests seen in euthyroid subjects, the epidemiology and mechanism of amiodarone-induced thyroid dysfunction, treatment options available, and the consequences of amiodarone use in pregnancy and lactation; and finally, we propose a follow-up strategy in patients taking amiodarone.

143

19477402

Which is the prognostic meaning of delayed enhancement documented in patients hypertrophic cardiomyopathy?

Delayed enhancement on cardiac magnetic resonance and clinical, morphological, and electrocardiographical features in hypertrophic cardiomyopathy. BACKGROUND: The clinical, morphological, and electrocardiographical relevance of delayed enhancement (DE) in cardiac magnetic resonance (CMR) was studied in patients with hypertrophic cardiomyopathy (HCM). METHODS AND RESULTS: A total of 56 patients underwent both gadolinium-enhanced CMR and 12-lead electrocardiogram. The CMR demonstrated DE at the left ventricular (LV) wall in 39 patients. The patients with DE included more cases with dilated phase of HCM, higher New York Heart Association (NYHA) classes and incidence of ventricular tachyarrhythmias (VT), lower LV ejection fraction (LVEF) and mean LV wall thickness (WT), and a larger ratio of maximum to minimum LVWT. The QRS duration was prolonged and the QRS axis deviated toward left with increases in the DE volume (r = 0.58 and r = 0.41, P < .01). Abnormal Q waves were present in 5 patients and the location coincided with the DE segments in 4 patients, but the concordance was not significant. The amplitude of T waves correlated with the ratio of the apex to basal LVWT (r = 0.38, P < .01) and was more negative in cases with DE at the apex. CONCLUSIONS: In HCM, the DE was associated with higher NYHA classes and prevalence of VT, impaired global LV function and asymmetrical hypertrophy, and conduction disturbance, abnormal Q waves, and giant negative T waves.

144

24861628

What is the COUGER tool?

COUGER--co-factors associated with uniquely-bound genomic regions. Most transcription factors (TFs) belong to protein families that share a common DNA binding domain and have very similar DNA binding preferences. However, many paralogous TFs (i.e. members of the same TF family) perform different regulatory functions and interact with different genomic regions in the cell. A potential mechanism for achieving this differential in vivo specificity is through interactions with protein co-factors. Computational tools for studying the genomic binding profiles of paralogous TFs and identifying their putative co-factors are currently lacking. Here, we present an interactive web implementation of COUGER, a classification-based framework for identifying protein co-factors that might provide specificity to paralogous TFs. COUGER takes as input two sets of genomic regions bound by paralogous TFs, and it identifies a small set of putative co-factors that best distinguish the two sets of sequences. To achieve this task, COUGER uses a classification approach, with features that reflect the DNA-binding specificities of the putative co-factors. The identified co-factors are presented in a user-friendly output page, together with information that allows the user to understand and to explore the contributions of individual co-factor features. COUGER can be run as a stand-alone tool or through a web interface: http://couger.oit.duke.edu.

145

24001228

Are there drugs for Tick-borne Encephalitis?

Current approaches in antiviral drug discovery against the Flaviviridae family. Viruses belonging to the Flaviviridae family primarily spread through arthropod vectors, and are the major causes of illness and death around the globe. The Flaviviridae family consists of 3 genera which include the Flavivirus genus (type species, yellow fever virus) as the largest genus, the Hepacivirus (type species, hepatitis C virus) and the Pestivirus (type species, bovine virus diarrhea). The flaviviruses (Flavivirus genus) are small RNA viruses transmitted by mosquitoes and ticks that take over host cell machinery in order to propagate. However, hepaciviruses and pestiviruses are not antropod-borne. Despite the extensive research and public health concern associated with flavivirus diseases, to date, there is no specific treatment available for any flavivirus infections, though commercially available vaccines for yellow fever, Japanese encephalitis and tick-born encephalitis exist. Due to the global threat of viral pandemics, there is an urgent need for new drugs. In many countries, patients with severe cases of flavivirus infections are treated only by supportive care, which includes intravenous fluids, hospitalization, respiratory support, and prevention of secondary infections. This review discusses the strategies used towards the discovery of antiviral drugs, focusing on rational drug design against Dengue virus (DENV), West Nile virus (WNV), Japanese encephalitis virus (JEV), Yellow Fever virus (YFV) and Hepatitis C virus (HCV). Only modified peptidic, nonpeptidic, natural compounds and fragment-based inhibitors (typically of mass less than 300 Da) against structural and non-structural proteins are discussed.

146

20402963

Is SLC22A3 expressed in the brain?

Interaction of organic cation transporter 3 (SLC22A3) and amphetamine. The organic cation transporter (OCT) 3 is widely expressed in various organs in humans, and involved in the disposition of many exogenous and endogenous compounds. Several lines of evidence have suggested that OCT3 expressed in the brain plays an important role in the regulation of neurotransmission. Relative to wild-type (WT) animals, Oct3 knockout (KO) mice have displayed altered behavioral and neurochemical responses to psychostimulants such as amphetamine (AMPH) and methamphetamine. In the present study, both in vitro and in vivo approaches were utilized to explore potential mechanisms underlying the disparate neuropharmacological effects observed following AMPH exposure in Oct3 KO mice. In vitro uptake studies conducted in OCT3 transfected cells indicated that dextroamphetamine (d-AMPH) is not a substrate of OCT3. However, OCT3 was determined to be a high-capacity and low-affinity transporter for the neurotransmitters dopamine (DA), norepinephrine (NE), and serotonin (5-HT). Inhibition studies demonstrated that d-AMPH exerts relatively weak inhibitory effects on the OCT3-mediated uptake of DA, NE, 5-HT, and the model OCT3 substrate 4-(4-(dimethylamino)styryl)-N-methylpyridinium iodide. The IC(50) values were determined to be 41.5 +/- 7.5 and 24.1 +/- 7.0 microM for inhibiting DA and 5-HT uptake, respectively, while 50% inhibition of NE and 4-(4-(dimethylamino)styryl)-N-methylpyridinium iodide uptake was not achieved by even the highest concentration of d-AMPH applied (100 microM). Furthermore, the disposition of d-AMPH in various tissues including the brain, liver, heart, kidney, muscle, intestine, spleen, testis, uterus, and plasma were determined in both male and female Oct3 KO and WT mice. No significant difference was observed between either genotypes or sex in all tested organs and tissues. Our findings suggest that OCT3 is not a prominent factor influencing the disposition of d-AMPH. Additionally, based upon the inhibitory potency observed in vitro, d-AMPH is unlikely to inhibit the uptake of monoamines mediated by OCT3 in the brain. Differentiated neuropharmacological effects of AMPHs noted between Oct3 KO and WT mice appear to be due to the absence of Oct3 mediated uptake of neurotransmitters in the KO mice.

147

20691807

Has the protein TIEG1 been associated with apoptosis?

The human papillomavirus-16 (HPV-16) oncoprotein E7 conjugates with and mediates the role of the transforming growth factor-beta inducible early gene 1 (TIEG1) in apoptosis. The human papillomavirus (HPV) oncoprotein E7 is a major transforming protein. The E7 protein does not possess intrinsic enzymatic activity, but rather functions through direct and indirect interactions with cellular proteins, several of which are well known cellular tumor suppressors. Using the yeast two-hybrid system, we found that transforming growth factor-beta inducible early gene 1 (TIEG1), a member of the Krüppel-like family (KLF) that has been implicated as a putative tumor suppressor, interacts and forms a specific complex with HPV-16 E7. TIEG1 has been shown to mimic the effects of TGF-beta in various carcinoma cells and plays a critical role in the apoptotic cascade. Our results indicate that E7 binds to the C-terminus of TIEG1 and induces its degradation via the ubiquitin pathway. E7 not only increased the ubiquitination of TIEG1 but also influenced the ability of TIEG1 to affect apoptosis. Our results suggest that suppression of TIEG1-mediated signaling by E7 may contribute to HPV-associated carcinogenesis.

148

25579226

Which is the cellular localization of the protein Opa1?

Release of mitochondrial Opa1 following oxidative stress in HT22 cells. Cellular mechanisms involved in multiple neurodegenerative diseases converge on mitochondria to induce overproduction of reactive oxygen species, damage to mitochondria, and subsequent cytochrome c release. Little is currently known regarding the contribution mitochondrial dynamics play in cytochrome c release following oxidative stress in neurodegenerative disease. Here we induced oxidative stress in the HT22 cell line with glutamate and investigated key mediators of mitochondrial dynamics to determine the role this process may play in oxidative stress induced neuronal death. We report that glutamate treatment in HT22 cells induces increase in reactive oxygen species (ROS), release of the mitochondrial fusion protein Opa1 into the cytosol, with concomitant release of cytochrome c. Furthermore, following the glutamate treatment alterations in cell signaling coincide with mitochondrial fragmentation which culminates in significant cell death in HT22 cells. Finally, we report that treatment with the antioxidant tocopherol attenuates glutamate induced-ROS increase, release of mitochondrial Opa1 and cytochrome c, and prevents cell death.

149

8725589

Which are the drugs utilized for the burning mouth syndrome?

Burning mouth syndrome. Burning mouth syndrome is a common condition particularly affecting elderly women. Numerous precipitating factors are recognized that lead to a burning sensation in clinically normal mucosa. By taking each precipitating factor into account, a favorable treatment outcome usually can be achieved. This article highlights the significance of precipitating factors in burning mouth syndrome and suggests a treatment protocol based on current scientific evidence.

150

11297621

Is PTEN involved in follicular thyroid carcinoma?

Somatic mutation and germline variants of MINPP1, a phosphatase gene located in proximity to PTEN on 10q23.3, in follicular thyroid carcinomas. Various genes have been identified to play a role in the pathogenesis of follicular thyroid tumors. Cowden syndrome is the only known familial syndrome with an increased risk of both follicular thyroid adenoma (FA) and carcinoma (FTC). Germline mutations in the tumor suppressor gene PTEN, which encodes a dual-specificity phosphatase, have been found in up to 80% of patients with Cowden syndrome suggesting a role of PTEN in the pathogenesis of follicular thyroid tumors. Although somatic intragenic mutations in PTEN, which maps to 10q23.3, are rarely found in follicular tumors, loss of heterozygosity (LOH) of markers within 10q22-24 occurs in about 25%. Recently, another phosphatase gene, MINPP1, has been localized to 10q23.3. MINPP1 has the ability to remove 3-phosphate from inositol phosphate substrates, a function that overlaps that of PTEN. Because of this overlapping function with PTEN and the physical location of MINPP1 to a region with frequent LOH in follicular thyroid tumors, we considered it to be an excellent candidate gene that could contribute to the pathogenesis of follicular thyroid tumors. We analyzed DNA from tumor and corresponding normal tissue from 23 patients with FA and 15 patients with FTC for LOH and mutations at the MINPP1 locus. LOH was identified in four malignant and three benign tumors. One of these FTCs with LOH was found to harbor a somatic c.122C > T or S41L mutation. We also found two germline sequence variants, c.809A > G (Q270R) and IVS3 + 34T > A. The c.809A > G variant was found in only one patient with FA but not in patients with FTC or normal controls. More interestingly, IVS3 + 34T > A was found in about 15% of FA cases and normal controls but not in patients with FTC. These results suggest a role for MINPP1 in the pathogenesis of at least a subset of malignant follicular thyroid tumors, and MINPP1 might act as a low penetrance predisposition allele for FTC.

151

25326329

Which genome browser database for DNA shape annotations is available?

GBshape: a genome browser database for DNA shape annotations. Many regulatory mechanisms require a high degree of specificity in protein-DNA binding. Nucleotide sequence does not provide an answer to the question of why a protein binds only to a small subset of the many putative binding sites in the genome that share the same core motif. Whereas higher-order effects, such as chromatin accessibility, cooperativity and cofactors, have been described, DNA shape recently gained attention as another feature that fine-tunes the DNA binding specificities of some transcription factor families. Our Genome Browser for DNA shape annotations (GBshape; freely available at http://rohslab.cmb.usc.edu/GBshape/) provides minor groove width, propeller twist, roll, helix twist and hydroxyl radical cleavage predictions for the entire genomes of 94 organisms. Additional genomes can easily be added using the GBshape framework. GBshape can be used to visualize DNA shape annotations qualitatively in a genome browser track format, and to download quantitative values of DNA shape features as a function of genomic position at nucleotide resolution. As biological applications, we illustrate the periodicity of DNA shape features that are present in nucleosome-occupied sequences from human, fly and worm, and we demonstrate structural similarities between transcription start sites in the genomes of four Drosophila species.

152

19008722

What is known about clinical efficacy of ceftriaxone for treatment of amyotrophic lateral sclerosis?

The beta-lactam antibiotic, ceftriaxone, dramatically improves survival, increases glutamate uptake and induces neurotrophins in stroke. OBJECTIVE: Ceftriaxone has been reported to reduce neuronal damage in amyotrophic lateral sclerosis and in an in-vitro model of neuronal ischaemia through increased expression and activity of the glutamate transporter, GLT1. We tested the effects of ceftriaxone on mortality, neurological outcome, and infarct size in experimental stroke in rats and looked for underlying mechanisms. METHODS: Male normotensive Wistar rats received ceftriaxone (200 mg/kg intraperitoneal) as a single injection 90 min after middle cerebral artery occlusion (90 min with reperfusion). Forty-eight hours after middle cerebral artery occlusion, infarct size (MRI) and neurological deficits were estimated. GLT1 expression was determined by real time RT-PCR, immunoblotting and promoter reporter assay, astrocyte GLT1 activity by measuring glutamate uptake. Bacterial load in various organs was measured by real time RT-PCR, neurotrophins and IL-6 by immunoblotting. RESULTS: Ceftriaxone dramatically reduced early (24-h) mortality from 34.5% (vehicle treatment, n = 29) to 0% (P < 0.01, n = 19). In a subgroup, followed up for 4 weeks, mortality persisted at 0%. Ceftriaxone strongly tended to reduce infarct size, it significantly improved neuronal survival within the penumbra, reduced neurological deficits (P < 0.001) and led to an upregulation of neurotrophins (P < 0.01) in the peri-infarct zone. Ceftriaxone did not increase GLT1 expression, but increased GLT1 activity (P < 0.05). CONCLUSION: Ceftriaxone causes a significant reduction in acute stroke mortality in a poststroke treatment regimen in animal studies. Improved neurological performance and survival may be due to neuroprotection by activation of GLT1 and a stimulation of neurotrophins resulting in an increased number of surviving neurons in the penumbra.

153

18922117

What is the relationship between TailorX and Oncotype?

Trial assessing individualized options for treatment for breast cancer: the TAILORx trial. Novel genetic profiling tests of breast cancer tissue have been shown to be prognostic for overall survival and predictive of local and distant rates of recurrence in breast cancer patients. One of these tests, Oncotype DXtrade mark, is a diagnostic test comprised of a 21-gene assay applied to paraffin-embedded breast cancer tissue, which allows physicians to predict subgroups of hormone-receptor-positive, node-negative patients who may benefit from hormonal therapy alone or require adjuvant chemotherapy to attain the best survival outcome. The results of the assay are converted to a recurrence score (0-100) that has been found to be predictive of 10- and 15-year local and distant recurrence in node-negative, estrogen-receptor-positive breast cancer patients. Previous studies have shown that patients with high recurrence scores benefit from adjuvant chemotherapy, whereas patients with low recurrence scores do not. To evaluate the ability to guide treatment decisions in the group with a mid-range recurrence score, the North American Cooperative Groups developed the Trial Assessing IndiviuaLized Options for Treatment for breast cancer, a randomized trial of chemotherapy followed by hormonal therapy versus hormonal therapy alone on invasive disease-free survival-ductal carcinoma in situ (IDFS-DCIS) survival in women with node-negative, estrogen-receptor-positive breast cancer with a recurrence score of 11-25. The study was initiated in May 2006 and approximately 4500 patients will be randomized. This article describes the rationale, methodology, statistical ana-lysis and implications of the results on clinical practice.

154

3101339

Does strenuous physical activity affect thyroid hormone metabolism?

Serum concentrations of thyrotropin, thyroxine, triiodothyronine and thyroxine binding globulin in female endurance runners and joggers. The effects of endurance training and season on the function of the anterior pituitary-thyroid axis were studied in 18 female runners and their 12 controls, and in 13 joggers and their 11 controls in Northern Finland, with a large seasonal difference in environmental factors. The serum concentrations of thyrotropin (TSH), thyroxine (T4), free thyroxine (fT4), triiodothyronine (T3), thyroxine binding globulin (TBG) and oestradiol (E2) were measured during one menstrual cycle in the light training season (autumn) and in the hard training season (spring). The responses of TSH to intravenous TRH stimulation were also measured in the luteal phase of the cycle during the hard training season. Endurance running did not affect the basal or TRH-stimulated serum TSH concentrations, while those of T4 and fT4 in runners were lowered in both seasons and that of T3 in the light training season in relation to control subjects. The serum concentrations of TBG were also significantly lower in runners than their controls in the luteal phase in both seasons. The effect of jogging on thyroid hormones was less pronounced. Serum concentrations of TSH, T4, fT4, T3 and TBG were generally slightly higher in spring than in autumn. Strenuous endurance training seems to have minor changes on the function of the thyroid gland. Depressed T4 levels in runners may rather be due to lowered TBG levels than due to direct effect of training. In spring the function of anterior pituitary-thyroid axis is more active than in autumn.

155

23446346

Which is the main function of "RNA sponges"?

Natural RNA circles function as efficient microRNA sponges. MicroRNAs (miRNAs) are important post-transcriptional regulators of gene expression that act by direct base pairing to target sites within untranslated regions of messenger RNAs. Recently, miRNA activity has been shown to be affected by the presence of miRNA sponge transcripts, the so-called competing endogenous RNA in humans and target mimicry in plants. We previously identified a highly expressed circular RNA (circRNA) in human and mouse brain. Here we show that this circRNA acts as a miR-7 sponge; we term this circular transcript ciRS-7 (circular RNA sponge for miR-7). ciRS-7 contains more than 70 selectively conserved miRNA target sites, and it is highly and widely associated with Argonaute (AGO) proteins in a miR-7-dependent manner. Although the circRNA is completely resistant to miRNA-mediated target destabilization, it strongly suppresses miR-7 activity, resulting in increased levels of miR-7 targets. In the mouse brain, we observe overlapping co-expression of ciRS-7 and miR-7, particularly in neocortical and hippocampal neurons, suggesting a high degree of endogenous interaction. We further show that the testis-specific circRNA, sex-determining region Y (Sry), serves as a miR-138 sponge, suggesting that miRNA sponge effects achieved by circRNA formation are a general phenomenon. This study serves as the first, to our knowledge, functional analysis of a naturally expressed circRNA.

156

22948232

Is the gene MAOA epigenetically modified by methylation?

Evidence that the methylation state of the monoamine oxidase A (MAOA) gene predicts brain activity of MAO A enzyme in healthy men. Human brain function is mediated by biochemical processes, many of which can be visualized and quantified by positron emission tomography (PET). PET brain imaging of monoamine oxidase A (MAO A)-an enzyme metabolizing neurotransmitters-revealed that MAO A levels vary widely between healthy men and this variability was not explained by the common MAOA genotype (VNTR genotype), suggesting that environmental factors, through epigenetic modifications, may mediate it. Here, we analyzed MAOA methylation in white blood cells (by bisulphite conversion of genomic DNA and subsequent sequencing of cloned DNA products) and measured brain MAO A levels (using PET and [(11)C]clorgyline, a radiotracer with specificity for MAO A) in 34 healthy non-smoking male volunteers. We found significant interindividual differences in methylation status and methylation patterns of the core MAOA promoter. The VNTR genotype did not influence the methylation status of the gene or brain MAO A activity. In contrast, we found a robust association of the regional and CpG site-specific methylation of the core MAOA promoter with brain MAO A levels. These results suggest that the methylation status of the MAOA promoter (detected in white blood cells) can reliably predict the brain endophenotype. Therefore, the status of MAOA methylation observed in healthy males merits consideration as a variable contributing to interindividual differences in behavior.

157

22137083

Which mutations of phopspholamban have been found in patients with cardiomyopathy?

Mutations in the human phospholamban gene in patients with heart failure. BACKGROUND: Phospholamban (PLN) is a crucial Ca(2+) cycling protein and a primary mediator of the β-adrenergic effects resulting in enhanced cardiac output. Mutations in the gene encoding PLN have been associated with idiopathic dilated cardiomyopathy; however, no systematic search for PLN mutations in heart failure has been conducted. METHODS: We screened a cohort of 1,014 Brazilian patients with heart failure for mutations in the PLN gene. Molecular modeling studies of the mutations found were developed. Different disease etiologies were present in our sample: idiopathic, ischemic, Chagas, valvular, hypertensive, and others. RESULTS: We identified 4 unrelated patients with PLN mutations (prevalence of 0.4%), 3 of them in the same amino acid residue (R9). Two patients presented a G-T missense mutation at the G26 nucleotide, which encodes an Arg-Leu substitution at codon 9 (R9L). One patient presented a G-A missense mutation at the same nucleotide, which encodes an Arg-His substitution at codon 9 (R9H). The fourth affected patient presented a T-G nonsense mutation at the nucleotide 116, substituting a termination codon for Leu-39 (L39stop). Molecular modeling studies suggested that R9L and R9H mutations might affect the region involved in protein kinase A docking and probably affect the mechanism modulating the release of phosphorylated PLN from the substrate binding site of protein kinase A. CONCLUSIONS: Mutations in the PLN gene are a rare cause of heart failure, present almost exclusively in patients with dilated cardiomyopathy etiology. The Arg9 and Leu39 residues are the leading location of mutations described at this locus to date. Despite the few mutated residues described to date, the clinical spectrum of presentation appears to vary considerably.

158

24323888

Which are the supplemental antioxidant in athletes?

Vitamin supplementation benefits in master athletes. Master athletes are more than 35 years of age and continue to train as hard as their young counterparts despite the aging process. All life long, they are capable of accomplishing exceptional sporting performances. For these participants in endurance events, matching energy intake and expenditure is critical to maintain health and performance. The proportions of carbohydrate, fat, and protein must be optimized to provide enough calories to sustain the energy requirements of competition or training, and for recovery. In addition, endurance athletes must include adequate vitamins and minerals in their diets to maintain healthy immune function. Vitamins and minerals may be sufficient in the diets of endurance athletes, who have a high energy intake. This would make it unnecessary to use vitamin and mineral supplements. Furthermore, one major limitation for these athletes is the management of oxidative stress, which, when in excess, can be deleterious for the organism. For individuals exposed to oxidative stress, micronutritional supplementations rich in vitamins and minerals can be also an alternative strategy. Although these supplementations are increasingly used by master athletes, very few data are available on their effects on oxidative stress, muscle recovery, and physical performance. The potential benefits of supplement use in athletes are thus questionable. Some studies indicate no benefits, while others highlight potential negative side effects of vitamin supplementation. Additional studies are warranted in order to design adapted prescriptions in antioxidant vitamins and minerals.

159

17035524

Is glycyl-tRNA synthetase gene involved in the development of Charcot-Marie-Tooth disease?

Functional analyses of glycyl-tRNA synthetase mutations suggest a key role for tRNA-charging enzymes in peripheral axons. Charcot-Marie-Tooth disease type 2D (CMT2D) and distal spinal muscular atrophy type V (dSMA-V) are axonal neuropathies characterized by a phenotype that is more severe in the upper extremities. We previously implicated mutations in the gene encoding glycyl-tRNA synthetase (GARS) as the cause of CMT2D and dSMA-V. GARS is a member of the family of aminoacyl-tRNA synthetases responsible for charging tRNA with cognate amino acids; GARS ligates glycine to tRNA(Gly). Here, we present functional analyses of disease-associated GARS mutations and show that there are not any significant mutation-associated changes in GARS expression levels; that the majority of identified GARS mutations modeled in yeast severely impair viability; and that, in most cases, mutant GARS protein mislocalizes in neuronal cells. Indeed, four of the five mutations studied show loss-of-function features in at least one assay, suggesting that tRNA-charging deficits play a role in disease pathogenesis. Finally, we detected endogenous GARS-associated granules in the neurite projections of cultured neurons and in the peripheral nerve axons of normal human tissue. These data are particularly important in light of the recent identification of CMT-associated mutations in another tRNA synthetase gene [YARS (tyrosyl-tRNA synthetase gene)]. Together, these findings suggest that tRNA-charging enzymes play a key role in maintaining peripheral axons.

160

21310746

Is there any software for automated analysis of FISH images?

FISH Finder: a high-throughput tool for analyzing FISH images. MOTIVATION: Fluorescence in situ hybridization (FISH) is used to study the organization and the positioning of specific DNA sequences within the cell nucleus. Analyzing the data from FISH images is a tedious process that invokes an element of subjectivity. Automated FISH image analysis offers savings in time as well as gaining the benefit of objective data analysis. While several FISH image analysis software tools have been developed, they often use a threshold-based segmentation algorithm for nucleus segmentation. As fluorescence signal intensities can vary significantly from experiment to experiment, from cell to cell, and within a cell, threshold-based segmentation is inflexible and often insufficient for automatic image analysis, leading to additional manual segmentation and potential subjective bias. To overcome these problems, we developed a graphical software tool called FISH Finder to automatically analyze FISH images that vary significantly. By posing the nucleus segmentation as a classification problem, compound Bayesian classifier is employed so that contextual information is utilized, resulting in reliable classification and boundary extraction. This makes it possible to analyze FISH images efficiently and objectively without adjustment of input parameters. Additionally, FISH Finder was designed to analyze the distances between differentially stained FISH probes. AVAILABILITY: FISH Finder is a standalone MATLAB application and platform independent software. The program is freely available from: http://code.google.com/p/fishfinder/downloads/list.

161

19571682

How do histone methyltransferases cause histone modification?

Degrees make all the difference: the multifunctionality of histone H4 lysine 20 methylation. Residue and degree-specific methylation of histone lysines along with other epigenetic modifications organizes chromatin into distinct domains and regulates almost every aspect of DNA metabolism. Identification of histone methyltransferases and demethylases, as well as proteins that recognize methylated lysines, has clarified the role of each methylation event in regulating different biological pathways. Methylation of histone H4 lysine 20 (H4K20me) plays critical roles in diverse cellular processes such as gene expression, cell cycle progression and DNA damage repair, with each of the three degrees of methylation (mono-, di- and tri-methylation) making a unique contribution. Here we discuss recent studies of H4K20me that have greatly improved our understanding of the regulation and function of this fascinating histone modification.

162

19327580

Is there an increased risk for cancer in Dyskeratosis Congenita?

Dyskeratosis congenita. Dyskeratosis congenita (DC) is an inherited bone marrow failure syndrome characterized clinically by the triad of abnormal nails, reticular skin pigmentation, and oral leukoplakia, and is associated with high risk of developing aplastic anemia, myelodysplastic syndrome, leukemia, and solid tumors. Patients have very short germline telomeres, and approximately half have mutations in one of six genes encoding proteins that maintain telomere function. Accurate diagnosis of DC is critical to ensure proper clinical management, because patients who have DC and bone marrow failure do not respond to immunosuppressive therapy and may have increased morbidity and mortality associated with hematopoietic stem cell transplantation.

163

19043405

Does MicroRNA-21 (miR-21) contribute to cardiovascular disease?

MicroRNA-21 contributes to myocardial disease by stimulating MAP kinase signalling in fibroblasts. MicroRNAs comprise a broad class of small non-coding RNAs that control expression of complementary target messenger RNAs. Dysregulation of microRNAs by several mechanisms has been described in various disease states including cardiac disease. Whereas previous studies of cardiac disease have focused on microRNAs that are primarily expressed in cardiomyocytes, the role of microRNAs expressed in other cell types of the heart is unclear. Here we show that microRNA-21 (miR-21, also known as Mirn21) regulates the ERK-MAP kinase signalling pathway in cardiac fibroblasts, which has impacts on global cardiac structure and function. miR-21 levels are increased selectively in fibroblasts of the failing heart, augmenting ERK-MAP kinase activity through inhibition of sprouty homologue 1 (Spry1). This mechanism regulates fibroblast survival and growth factor secretion, apparently controlling the extent of interstitial fibrosis and cardiac hypertrophy. In vivo silencing of miR-21 by a specific antagomir in a mouse pressure-overload-induced disease model reduces cardiac ERK-MAP kinase activity, inhibits interstitial fibrosis and attenuates cardiac dysfunction. These findings reveal that microRNAs can contribute to myocardial disease by an effect in cardiac fibroblasts. Our results validate miR-21 as a disease target in heart failure and establish the therapeutic efficacy of microRNA therapeutic intervention in a cardiovascular disease setting.

164

20681793

What is the enzymatic activity of the breast cancer associated gene BRCA1?

Impact of RING and BRCT domain mutations on BRCA1 protein stability, localization and recruitment to DNA damage. Mutations within the tumor suppressor BRCA1 cause the majority of hereditary breast and ovarian cancers. The BRCA1 protein is an important regulator of DNA double-strand break repair, and BRCA1-deficient cells are highly sensitive to ionizing radiation. Furthermore, BRCA1 function may contribute to enforcement of the G(2) cell cycle checkpoint. E3-ubiquitin ligase activity is the only known enzymatic activity of BRCA1, which is mediated by the N-terminal RING finger domain. The C-terminal BRCT repeat domain, which mediates protein-protein interactions, is the only other identified structural domain. By investigating cancer-linked mutations within each domain, we demonstrate that truncation of the BRCT domain greatly impairs the stability and nuclear localization of BRCA1 protein. A missense mutation within the RING domain does not affect these biochemical properties. However, both mutant forms of BRCA1 fail to colocalize in nuclear foci with the known BRCA1-interacting proteins BARD1 and BACH1, which are important for DNA repair. This failure occurs despite the continued ability of the RING mutant protein to interact with BACH1 and the ability of the BRCT mutant to interact with BARD1. Furthermore, neither mutant form of BRCA1 is recruited into DNA damage-associated foci marked by gamma-H2AX. Therefore, our data suggest that both the RING and BRCT domains of BRCA1 are required for an early step in the function of BRCA1 during DNA repair: recruitment to the sites of DNA damage.

165

22807527

List markers for autophagy.

Chlamydia trachomatis vacuole maturation in infected macrophages. Chlamydia trachomatis is an obligate intracellular bacterium responsible for one of the most common sexually transmitted diseases. In epithelial cells, C. trachomatis resides in a modified membrane-bound vacuole known as an inclusion, which is isolated from the endocytic pathway. However, the maturation process of C. trachomatis within immune cells, such as macrophages, has not been studied extensively. Here, we demonstrated that RAW macrophages effectively suppressed C. trachomatis growth and prevented Golgi stack disruption, a hallmark defect in epithelial cells after C. trachomatis infection. Next, we systematically examined association between C. trachomatis and various endocytic pathway markers. Spinning disk confocal time-lapse studies revealed significant and rapid association between C. trachomatis with Rab7 and LAMP1, markers of late endosomes and lysosomes. Moreover, pretreatment with an inhibitor of lysosome acidification led to significant increases in C. trachomatis growth in macrophages. At later stages of infection, C. trachomatis associated with the autophagy marker LC3. TEM analysis confirmed that a significant portion of C. trachomatis resided within double-membrane-bound compartments, characteristic of autophagosomes. Together, these results suggest that macrophages can suppress C. trachomatis growth by targeting it rapidly to lysosomes; moreover, autophagy is activated at later stages of infection and targets significant numbers of the invading bacteria, which may enhance subsequent chlamydial antigen presentation.

166

22870189

Are there any statistical methods for normalizing and identifying differential regions in histone modification ChIP-seq data?

ChIPnorm: a statistical method for normalizing and identifying differential regions in histone modification ChIP-seq libraries. The advent of high-throughput technologies such as ChIP-seq has made possible the study of histone modifications. A problem of particular interest is the identification of regions of the genome where different cell types from the same organism exhibit different patterns of histone enrichment. This problem turns out to be surprisingly difficult, even in simple pairwise comparisons, because of the significant level of noise in ChIP-seq data. In this paper we propose a two-stage statistical method, called ChIPnorm, to normalize ChIP-seq data, and to find differential regions in the genome, given two libraries of histone modifications of different cell types. We show that the ChIPnorm method removes most of the noise and bias in the data and outperforms other normalization methods. We correlate the histone marks with gene expression data and confirm that histone modifications H3K27me3 and H3K4me3 act as respectively a repressor and an activator of genes. Compared to what was previously reported in the literature, we find that a substantially higher fraction of bivalent marks in ES cells for H3K27me3 and H3K4me3 move into a K27-only state. We find that most of the promoter regions in protein-coding genes have differential histone-modification sites. The software for this work can be downloaded from http://lcbb.epfl.ch/software.html.

167

21094032

Is CD84 genetically associated with arthritis?

The role of SLAM/CD2 polymorphisms in systemic autoimmunity. The SLAM/CD2 gene family encodes receptors that play important roles in regulating multiple cellular interactions in the adaptive and innate immune systems. Three members of this gene family, Ly108, Ly9, and CD84, exhibit polymorphisms that strongly influence susceptibility to systemic autoimmunity, notably in mice, but also in some human populations. Polymorphisms of Ly108 in mice strongly impact central tolerance in both B and T cell development, predominantly by modulating apoptosis, anergy, and cell-cycle progression. In addition, Ly108 and CD84, together with their downstream signaling adaptor SLAM-associated protein (SAP), have emerged as key players in B-T interactions during the formation of germinal centers. Interestingly, several independent lines of research have now associated variations in B-T interactions during germinal center formation with systemic autoimmunity, suggesting that susceptibility to systemic lupus erythematosus (SLE) may involve in part the impairment of this peripheral tolerance checkpoint. These new insights into the multiplicity of roles played by the SLAM/CD2 family and its potential importance in human autoimmunity positions the SLAM/CD2 family as an excellent target for immunotherapy.

168

25003133

What is the function of Neu5Gc (N-Glycolylneuraminic acid)?

Prevalence of anti-Neu5Gc antibodies in patients with hypothyroidism. BACKGROUND: N-Glycolylneuraminic acid (Neu5Gc) is a sialic acid synthesized by animals, but not by humans or birds. However, it can be incorporated in human cells and can trigger immune response. In the present study, we detected anti-Neu5Gc antibodies in samples of the general population and of patients suffering from hypothyroidism/Hashimoto's disease, which is known to have autoimmune origin. METHODS: Antibodies were measured using enzyme-immunosorbent techniques. RESULTS: Serum anti-Neu5Gc IgG antibodies were higher in patients with hypothyroidism (mean: 14.8 ± 15.9 μg/mL, median: 10.0 μg/mL, P = 0.0003, Mann-Whitney) and even higher in the group with Hashimoto's thyroiditis (mean: 31.1 ± 16.3 μg/mL, median: 27.2 μg/mL, P = 0.0000, Mann-Whitney) compared to the general population (mean: 5.3 ± 4.7 μg/mL, median : 4 μg/mL). All anti-TPO positive samples had anti-Neu5Gc antibody concentration higher than the mean value of the general population while anti-TPO concentration was increased as anti-Neu5Gc concentration increased. Low concentrations of IgA and IgM antibodies were measured in both general population and patient groups. CONCLUSION: The increased values of anti-Neu5Gc antibodies in patients with hypothyroidism/Hashimoto's disease and the correlation of anti-TPO incidence with increased anti-Neu5Gc concentration raise the possibility of an association between anti-Neu5Gc antibody development and autoimmune hypothyroidism.

169

12871541

Are there any specific antidotes for rivaroxaban?

The use of specific antidotes as a response to bleeding complications during anticoagulant therapy for venous thromboembolism. When a bleeding complication occurs during therapy with heparin or vitamin K antagonists, there is an option to give a specific antidote. Several new anticoagulants have been developed that are likely to have some risk of bleeding complications, for which no specific antidotes are available. Interestingly, it is unknown how often the use of an antidote is necessary in clinical practice. We investigated 1877 patients treated for venous thromboembolism included in three large clinical trials, of which 181 (9.6%) had a total of 225 adjudicated bleeding episodes; 46 hemorrhages being designated as major. Some form of antidote was given to 26 (14.4%) patients with a hemorrhage. Of the patients with at least one major hemorrhage, 19 (41.3%) received an antidote. Vitamin K was given to 23 (1.2%) patients, one (0.05%) patient received protamin sulfate and seven (0.4%) patients received fresh frozen plasma. The use of antidotes was comparable for initial and long-term treatment. Antidotes were statistically significantly more frequently given in Canada as compared to other participating countries. Vitamin K was more frequently given in case of a higher international normalized ratio value. Although antidotes against anticoagulant treatment are widely available, our analysis shows that in only a very small number of patients a direct, or slow-acting antidote to reverse the anticoagulant effect was used.

170

9106214

Which metabolite activates AtxA?

Cross-talk to the genes for Bacillus anthracis capsule synthesis by atxA, the gene encoding the trans-activator of anthrax toxin synthesis. The two major virulence factors of Bacillus anthracis are the tripartite toxin and the polyglutamate capsule, which are encoded by genes on the large plasmids, pXO1 and pXO2, respectively. The genes atxA, located on pXO1, and acpA, located on pXO2, encode positive trans-acting proteins that are involved in bicarbonate-mediated regulation of toxin and capsule production, respectively. A derivative strain cured of pXO1 produced less capsular substance than the parent strain harbouring both pXO1 and pXO2, and electroporation of the strain cured of pXO1 with a plasmid containing the cloned atxA gene resulted in an increased level of capsule production. An acpA-null mutant was complemented by not only acpA but also the atxA gene. The cap region, which is essential for encapsulation, contains three genes capB, capC, and capA, arranged in that order. The atxA gene stimulated capsule synthesis from the cloned cap region. Transcriptional analysis of cap by RNA slot-blot hybridization and primer-extension analysis revealed that atxA activated expression of cap in trans at the transcriptional level. These results indicate that cross-talk occurs, in which the pXO1-located gene, atxA, activates transcription of the cap region genes located on pXO2. We identified two major apparent transcriptional start sites, designated P1 and P2, located at positions 731 bp and 625 bp, respectively, upstream of the translation-initiation codon of capB. Transcription initiated from P1 and P2 was activated by both atxA and acpA, and activation appeared to be stimulated by bicarbonate. Deletion analysis of the upstream region of the cap promoter revealed that activation by both atxA and acpA required a DNA segment of 70 bp extending upstream of the P1 site. These results suggest that cross-talk by atxA to the genes encoding capsule synthesis is caused by the interaction of the atxA gene product with a regulatory sequence upstream of cap.

171

58611

What is the function of 6SRNA in bacteria?

Structure and function of RNA replicase of bacteriophage Qbeta. (1) The RNA replicase induced by bacteriophage Qbeta consists of four non-identical subunits designated as alpha (mol. wt. 74000), beta (mol. wt. 64000), gamma (mol. wt. 47000) and delta (mol. wt. 33000), only one (subunit beta) of which is specified by the phage genome. (2) Subunit alpha (30 S ribosomal protein "S1" as well as translational interference factor "i") is required only for (+) strand-directed RNA synthesis in the presence of the host factor. (3) Qbeta replicase lacking subunit alpha (R-alpha) is capable of replicating templates other than (+) strand, such as (--), "6S" RNA, poly(C) etc., in the absence of the host factor. (4) Subunit beta is suggested to be the nucleotide-polymerizing enzyme, but is unable to initiate RNA synthesis by itself. (5) Subunits gamma and delta are identical to the protein synthesis elongation factors, EF-Tu and EF-Ts, respectively, and are required only for initiation of RNA synthesis, but not for elongation. (6) A model of Qbeta replicase is presented in order to discuss observed template-enzyme interactions.

172

22513936

Is cytisine superior to nicotine replacement therapy for smoking cessation?

Nicotine receptor partial agonists for smoking cessation. BACKGROUND: Nicotine receptor partial agonists may help people to stop smoking by a combination of maintaining moderate levels of dopamine to counteract withdrawal symptoms (acting as an agonist) and reducing smoking satisfaction (acting as an antagonist). OBJECTIVES: The primary objective of this review is to assess the efficacy and tolerability of nicotine receptor partial agonists, including cytisine, dianicline and varenicline for smoking cessation. SEARCH METHODS: We searched the Cochrane Tobacco Addiction Group's specialised register for trials, using the terms ('cytisine' or 'Tabex' or 'dianicline' or 'varenicline' or 'nicotine receptor partial agonist') in the title or abstract, or as keywords. The register is compiled from searches of MEDLINE, EMBASE, PsycINFO and Web of Science using MeSH terms and free text to identify controlled trials of interventions for smoking cessation and prevention. We contacted authors of trial reports for additional information where necessary. The latest update of the specialised register was in December 2011. We also searched online clinical trials registers. SELECTION CRITERIA: We included randomized controlled trials which compared the treatment drug with placebo. We also included comparisons with bupropion and nicotine patches where available. We excluded trials which did not report a minimum follow-up period of six months from start of treatment. DATA COLLECTION AND ANALYSIS: We extracted data on the type of participants, the dose and duration of treatment, the outcome measures, the randomization procedure, concealment of allocation, and completeness of follow-up.The main outcome measured was abstinence from smoking at longest follow-up. We used the most rigorous definition of abstinence, and preferred biochemically validated rates where they were reported. Where appropriate we pooled risk ratios (RRs), using the Mantel-Haenszel fixed-effect model. MAIN RESULTS: Two recent cytisine trials (937 people) found that more participants taking cytisine stopped smoking compared with placebo at longest follow-up, with a pooled RR of 3.98 (95% confidence interval (CI) 2.01 to 7.87). One trial of dianicline (602 people) failed to find evidence that it was effective (RR 1.20, 95% CI 0.82 to 1.75). Fifteen trials compared varenicline with placebo for smoking cessation; three of these also included a bupropion treatment arm. We also found one open-label trial comparing varenicline plus counselling with counselling alone. We found one relapse prevention trial, comparing varenicline with placebo, and two open-label trials comparing varenicline with nicotine replacement therapy (NRT). We also include one trial in which all the participants were given varenicline, but received behavioural support either online or by phone calls, or by both methods. This trial is not included in the analyses, but contributes to the data on safety and tolerability. The included studies covered 12,223 participants, 8100 of whom used varenicline.The pooled RR for continuous or sustained abstinence at six months or longer for varenicline at standard dosage versus placebo was 2.27 (95% CI 2.02 to 2.55; 14 trials, 6166 people, excluding one trial evaluating long term safety). Varenicline at lower or variable doses was also shown to be effective, with an RR of 2.09 (95% CI 1.56 to 2.78; 4 trials, 1272 people). The pooled RR for varenicline versus bupropion at one year was 1.52 (95% CI 1.22 to 1.88; 3 trials, 1622 people). The RR for varenicline versus NRT for point prevalence abstinence at 24 weeks was 1.13 (95% CI 0.94 to 1.35; 2 trials, 778 people). The two trials which tested the use of varenicline beyond the 12-week standard regimen found the drug to be well-tolerated during long-term use. The main adverse effect of varenicline was nausea, which was mostly at mild to moderate levels and usually subsided over time. A meta-analysis of reported serious adverse events occurring during or after active treatment and not necessarily considered attributable to treatment suggests there may be a one-third increase in the chance of severe adverse effects among people using varenicline (RR 1.36; 95% CI 1.04 to 1.79; 17 trials, 7725 people), but this finding needs to be tested further. Post-marketing safety data have raised questions about a possible association between varenicline and depressed mood, agitation, and suicidal behaviour or ideation. The labelling of varenicline was amended in 2008, and the manufacturers produced a Medication Guide. Thus far, surveillance reports and secondary analyses of trial data are inconclusive, but the possibility of a link between varenicline and serious psychiatric or cardiovascular events cannot be ruled out. AUTHORS' CONCLUSIONS: Cytisine increases the chances of quitting, although absolute quit rates were modest in two recent trials. Varenicline at standard dose increased the chances of successful long-term smoking cessation between two- and threefold compared with pharmacologically unassisted quit attempts. Lower dose regimens also conferred benefits for cessation, while reducing the incidence of adverse events. More participants quit successfully with varenicline than with bupropion. Two open-label trials of varenicline versus NRT suggested a modest benefit of varenicline but confidence intervals did not rule out equivalence. Limited evidence suggests that varenicline may have a role to play in relapse prevention. The main adverse effect of varenicline is nausea, but mostly at mild to moderate levels and tending to subside over time. Possible links with serious adverse events, including serious psychiatric or cardiovascular events, cannot be ruled out.Future trials of cytisine may test extended regimens and more intensive behavioural support. There is a need for further trials of the efficacy of varenicline treatment extended beyond 12 weeks.

173

24086431

Which amino acid residue appears mutated in most of the cases reported with cadasil syndrome?

NOTCH3 variants and risk of ischemic stroke. BACKGROUND: Mutations within the NOTCH3 gene cause cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL). CADASIL mutations appear to be restricted to the first twenty-four exons, resulting in the gain or loss of a cysteine amino acid. The role of other exonic NOTCH3 variation not involving cysteine residues and mutations in exons 25-33 in ischemic stroke remains unresolved. METHODS: All 33 exons of NOTCH3 were sequenced in 269 Caucasian probands from the Siblings With Ischemic Stroke Study (SWISS), a 70-center North American affected sibling pair study and 95 healthy Caucasian control subjects. Variants identified by sequencing in the SWISS probands were then tested for association with ischemic stroke using US Caucasian controls collected at the Mayo Clinic (n=654), and further assessed in a Caucasian (n=802) and African American (n=298) patient-control series collected through the Ischemic Stroke Genetics Study (ISGS). RESULTS: Sequencing of the 269 SWISS probands identified one (0.4%) with small vessel type stroke carrying a known CADASIL mutation (p.R558C; Exon 11). Of the 19 common NOTCH3 variants identified, the only variant significantly associated with ischemic stroke after multiple testing adjustment was p.R1560P (rs78501403; Exon 25) in the combined SWISS and ISGS Caucasian series (Odds Ratio [OR] 0.50, P=0.0022) where presence of the minor allele was protective against ischemic stroke. Although only significant prior to adjustment for multiple testing, p.T101T (rs3815188; Exon 3) was associated with an increased risk of small-vessel stroke (OR: 1.56, P=0.008) and p.P380P (rs61749020; Exon 7) was associated with decreased risk of large-vessel stroke (OR: 0.35, P=0.047) in Caucasians. No significant associations were observed in the small African American series. CONCLUSION: Cysteine-affecting NOTCH3 mutations are rare in patients with typical ischemic stroke, however our observation that common NOTCH3 variants may be associated with risk of ischemic stroke warrants further study.

174

21856302

Which syndromes are associated with mutations in the EZH2 gene?

The Y641C mutation of EZH2 alters substrate specificity for histone H3 lysine 27 methylation states. Mutations at tyrosine 641 (Y641F, Y641N, Y641S and Y641H) in the SET domain of EZH2 have been identified in patients with certain subtypes of non-Hodgkin lymphoma (NHL). These mutations were shown to change the substrate specificity of EZH2 for various methylation states of lysine 27 on histone H3 (H3K27). An additional mutation at EZH2 Y641 to cysteine (Y641C) was also found in one patient with NHL and in SKM-1 cells derived from a patient with myelodisplastic syndrome (MDS). The Y641C mutation has been reported to dramatically reduce enzymatic activity. Here, we demonstrate that while the Y641C mutation ablates enzymatic activity against unmethylated and monomethylated H3K27, it is superior to wild-type in catalyzing the formation of trimethylated H3K27 from the dimethylated precursor.

175

12520035

Which databases exist for experimentally determined topologies of α-helical transmembrane proteins ?

TMPDB: a database of experimentally-characterized transmembrane topologies. TMPDB is a database of experimentally-characterized transmembrane (TM) topologies. TMPDB release 6.2 contains a total of 302 TM protein sequences, in which 276 are alpha-helical sequences, 17 beta-stranded, and 9 alpha-helical sequences with short pore-forming helices buried in the membrane. The TM topologies in TMPDB were determined experimentally by means of X-ray crystallography, NMR, gene fusion technique, substituted cysteine accessibility method, N-linked glycosylation experiment and other biochemical methods. TMPDB would be useful as a test and/or training dataset in improving the proposed TM topology prediction methods or developing novel methods with higher performance, and as a guide for both the bioinformaticians and biologists to better understand TM proteins. TMPDB and its subsets are freely available at the following web site: http://bioinfo.si.hirosaki-u.ac.jp/~TMPDB/.

176

23272425

Which disease is characterized by congenital absence of intrinsic ganglion cells of the gastrointestinal tract?

Morphological patterns in children with ganglion related enteric neuronal abnormalities. BACKGROUND: Hirschsprung's Disease (HD) is a developmental disorder of enteric nervous system characterised by the absence of ganglion cells in submucosal (Meissner's) and myenteric (Aurbach's) plexuses of distal bowel. The purpose of the present study was to observe and report the morphological patterns of ganglion related enteric neuronal abnormalities in children presented with clinical features of (HD) in a Pakistani population. METHODS: A total of 92 patients with clinical presentation of HD were enrolled between March 2009 and October 2009. Among them, 8 were excluded according to the exclusion criteria. After detailed history and physical examination, paraffin embedded H and E stained sections were prepared from the serial open biopsies from colorectum. The data was analysed using SPSS-17. Frequencies and percentages are given for qualitative variables. Non-parametric Binomial Chi-Square test was applied to observe within group associations and p<0.05 was considered statistically significant. RESULTS: Among 84 patients, 13 (15.5%) proved to be normally ganglionic whereas 71 (84.5%) showed ganglion related enteric neuronal abnormalities namely isolated hypoganglionosis 9 (12.7%), immaturity of ganglion cells 9 (12.7%), isolated hyperganglionosis (IND Type B) 2 (2.8%) and Hirschsprung's disease 51 (71.8%). Among HD group, 34 (66.7%) belonged to isolated form and 17 (33.3%) showed combined ganglion related abnormalities. CONCLUSIONS: Hirschsprung's disease is common in Pakistani population, followed by hypoganglionosis, immaturity of ganglion cells and IND type B. The presence of hypertrophic nerve fibres was significant in HD, hyperganglionosis and hypoganglionosis, whereas, no hypertrophic nerve fibres were appreciated in immaturity of ganglion cell group.

177

20132664

What is the disease in which patients are sensitive to DNA crosslinking agents, presenting with a high frequency of chromosomal aberrations?

Clinical, genetic and cytogenetic study of Fanconi anemia in an Indian population. Fanconi anemia (FA) is a rare autosomal recessive genetic disease, associated with congenital anomalies and a predisposition to cancers. FA patients exhibit spontaneous chromosome breakage and FA cells are sensitive to DNA interstrand crosslink agents and expresses high frequency of chromosome breakage. Recently 13 genes have been shown to be involved with the FA phenotype. We have carried out a detailed study in clinically diagnosed FA patients in an Indian population. Thirty three patients were clinically diagnosed with FA and had aplastic anemia and bleeding abnormalities. The genetic analysis revealed a significantly (P<0.0001) high frequency (36.4%) of parental consanguinity in FA patients compared to controls (3.33%). Chromosomal analysis revealed spontaneous chromosome breakage in 63.64% FA patients. The mitomycin C and diepoxybutane induced cultures showed a significantly (P<0.001) high frequency of chromosome breakage and radial formation compared to controls. Among 33 patients, nine (27.27%) patients developed malignancies and chromosomal abnormalities were detected in five (55.5%) patients bone marrow cells including monosomy 5 and 7, trisomy 10, der(1q) and inv(7). Cytogenetic investigation is important in aplastic anemia to rule out FA. The clinical presentation and the associated high frequency of consanguinity in FA, and the molecular analysis are complementary in the study of an Indian population.

178

24915039

How is oprozomib administered?

Carfilzomib and oprozomib synergize with histone deacetylase inhibitors in head and neck squamous cell carcinoma models of acquired resistance to proteasome inhibitors. Acquired resistance to proteasome inhibitors represents a considerable impediment to their effective clinical application. Carfilzomib and its orally bioavailable structural analog oprozomib are second-generation, highly-selective, proteasome inhibitors. However, the mechanisms of acquired resistance to carfilzomib and oprozomib are incompletely understood, and effective strategies for overcoming this resistance are needed. Here, we developed models of acquired resistance to carfilzomib in two head and neck squamous cell carcinoma cell lines, UMSCC-1 and Cal33, through gradual exposure to increasing drug concentrations. The resistant lines R-UMSCC-1 and R-Cal33 demonstrated 205- and 64-fold resistance, respectively, relative to the parental lines. Similarly, a high level of cross-resistance to oprozomib, as well as paclitaxel, was observed, whereas only moderate resistance to bortezomib (8- to 29-fold), and low level resistance to cisplatin (1.5- to 5-fold) was seen. Synergistic induction of apoptosis signaling and cell death, and inhibition of colony formation followed co-treatment of acquired resistance models with carfilzomib and the histone deacetylase inhibitor (HDACi) vorinostat. Synergism was also seen with other combinations, including oprozomib plus vorinostat, or carfilzomib plus the HDACi entinostat. Synergism was accompanied by upregulation of proapoptotic Bik, and suppression of Bik attenuated the synergy. The acquired resistance models also exhibited elevated levels of MDR-1/P-gp. Inhibition of MDR-1/P-gp with reversin 121 partially overcame carfilzomib resistance in R-UMSCC-1 and R-Cal33 cells. Collectively, these studies indicate that combining carfilzomib or oprozomib with HDAC or MDR-1/P-gp inhibitors may be a useful strategy for overcoming acquired resistance to these proteasome inhibitors.

179

24013648

What is a Caveolae?

Molecular composition and ultrastructure of the caveolar coat complex. Caveolae are an abundant feature of the plasma membrane of many mammalian cell types, and have key roles in mechano-transduction, metabolic regulation, and vascular permeability. Caveolin and cavin proteins, as well as EHD2 and pacsin 2, are all present in caveolae. How these proteins assemble to form a protein interaction network for caveolar morphogenesis is not known. Using in vivo crosslinking, velocity gradient centrifugation, immuno-isolation, and tandem mass spectrometry, we determine that cavins and caveolins assemble into a homogenous 80S complex, which we term the caveolar coat complex. There are no further abundant components within this complex, and the complex excludes EHD2 and pacsin 2. Cavin 1 forms trimers and interacts with caveolin 1 with a molar ratio of about 1∶4. Cavins 2 and 3 compete for binding sites within the overall coat complex, and form distinct subcomplexes with cavin 1. The core interactions between caveolin 1 and cavin 1 are independent of cavin 2, cavin 3, and EHD2 expression, and the cavins themselves can still interact in the absence of caveolin 1. Using immuno-electron microscopy as well as a recently developed protein tag for electron microscopy (MiniSOG), we demonstrate that caveolar coat complexes form a distinct coat all around the caveolar bulb. In contrast, and consistent with our biochemical data, EHD2 defines a different domain at the caveolar neck. 3D electron tomograms of the caveolar coat, labeled using cavin-MiniSOG, show that the caveolar coat is composed of repeating units of a unitary caveolar coat complex.

180

15791412

Which are the roles of chromatin compartments in the eukaryotic nucleus?

Plasticity of histone modifications across the invertebrate to vertebrate transition: histone H3 lysine 4 trimethylation in heterochromatin. Histone posttranslational modifications mediate establishment of structurally and functionally distinct chromatin compartments of eukaryotic nuclei. The association of different histone modifications with euchromatic and heterochromatic compartments is relatively conserved in highly divergent model organisms such as Drosophila and mammals. However, some differences between these model systems have been uncovered while limited data are available from organisms nearer the invertebrate-vertebrate transition. We identified a chromatin compartment in both diploid and endocycling cells of the urochordate, Oikopleura dioica, enriched in heterochromatic histone modifications and DNA methylation. Surprisingly, this compartment also contained high levels of histone H3 trimethylated at lysine 4 (H3 Me(3)K4), a modification thus far associated with actively transcribed sequences. Although in Drosophila and mouse cells, H3 Me(3)K4 was prevalently associated with euchromatin, we also detected it in their pericentromeric heterochromatin. We further showed that H3 Me(3)K4 abundance was not necessarily proportional to local levels of transcriptional activity in either euchromatin or heterochromatin. Our data indicate greater plasticity across evolution in the association of histone lysine methylation with functionally distinct chromatin domains than previously thought and suggest that H3 Me(3)K4 participates in additional processes beyond marking transcriptionally active chromatin.

181

25340765

Is the abnormal dosage of ultraconserved elements disfavored in cancer cells?

Abnormal dosage of ultraconserved elements is highly disfavored in healthy cells but not cancer cells. Ultraconserved elements (UCEs) are strongly depleted from segmental duplications and copy number variations (CNVs) in the human genome, suggesting that deletion or duplication of a UCE can be deleterious to the mammalian cell. Here we address the process by which CNVs become depleted of UCEs. We begin by showing that depletion for UCEs characterizes the most recent large-scale human CNV datasets and then find that even newly formed de novo CNVs, which have passed through meiosis at most once, are significantly depleted for UCEs. In striking contrast, CNVs arising specifically in cancer cells are, as a rule, not depleted for UCEs and can even become significantly enriched. This observation raises the possibility that CNVs that arise somatically and are relatively newly formed are less likely to have established a CNV profile that is depleted for UCEs. Alternatively, lack of depletion for UCEs from cancer CNVs may reflect the diseased state. In support of this latter explanation, somatic CNVs that are not associated with disease are depleted for UCEs. Finally, we show that it is possible to observe the CNVs of induced pluripotent stem (iPS) cells become depleted of UCEs over time, suggesting that depletion may be established through selection against UCE-disrupting CNVs without the requirement for meiotic divisions.

182

19298522

Does thyroid hormone regulate calcium transient in the myocardium?

Modulation of cardiac ionic homeostasis by 3-iodothyronamine. 3-iodothyronamine (T(1)AM) is a novel endogenous relative of thyroid hormone, able to interact with trace amine-associated receptors, a class of plasma membrane G protein-coupled receptors, and to produce a negative inotropic and chronotropic effect. In the isolated rat heart 20-25 microM T(1)AM decreased cardiac contractility, but oxygen consumption and glucose uptake were either unchanged or disproportionately high when compared to mechanical work. In adult rat cardiomyocytes acute exposure to 20 microM T(1)AM decreased the amplitude and duration of the calcium transient. In patch clamped cardiomyocytes sarcolemmal calcium current density was unchanged while current facilitation by membrane depolarization was abolished consistent with reduced sarcoplasmic reticulum (SR) calcium release. In addition, T(1)AM decreased transient outward current (I(to)) and I(K1) background current. SR studies involving 20 microM T(1)AM revealed a significant decrease in ryanodine binding due to reduced B(max), no significant change in the rate constant of calcium-induced calcium release, a significant increase in calcium leak measured under conditions promoting channel closure, and no effect on oxalate-supported calcium uptake. Based on these observations we conclude T(1)AM affects calcium and potassium homeostasis and suggest its negative inotropic action is due to a diminished pool of SR calcium as a result of increased diastolic leak through the ryanodine receptor, while increased action potential duration is accounted for by inhibition of I(to) and I(K1) currents.

183

22337885

What are the biological roles proposed for proteins containing the SPRY domain?

TRIM67 protein negatively regulates Ras activity through degradation of 80K-H and induces neuritogenesis. Tripartite motif (TRIM)-containing proteins, which are defined by the presence of a common domain structure composed of a RING finger, one or two B-box motifs and a coiled-coil motif, are involved in many biological processes including innate immunity, viral infection, carcinogenesis, and development. Here we show that TRIM67, which has a TRIM motif, an FN3 domain and a SPRY domain, is highly expressed in the cerebellum and that TRIM67 interacts with PRG-1 and 80K-H, which is involved in the Ras-mediated signaling pathway. Ectopic expression of TRIM67 results in degradation of endogenous 80K-H and attenuation of cell proliferation and enhances neuritogenesis in the neuroblastoma cell line N1E-115. Furthermore, morphological and biological changes caused by knockdown of 80K-H are similar to those observed by overexpression of TRIM67. These findings suggest that TRIM67 regulates Ras signaling via degradation of 80K-H, leading to neural differentiation including neuritogenesis.

184

19582169

How could we infer functional associations from gene fusion events?

Multiple horizontal gene transfer events and domain fusions have created novel regulatory and metabolic networks in the oomycete genome. Complex enzymes with multiple catalytic activities are hypothesized to have evolved from more primitive precursors. Global analysis of the Phytophthora sojae genome using conservative criteria for evaluation of complex proteins identified 273 novel multifunctional proteins that were also conserved in P. ramorum. Each of these proteins contains combinations of protein motifs that are not present in bacterial, plant, animal, or fungal genomes. A subset of these proteins were also identified in the two diatom genomes, but the majority of these proteins have formed after the split between diatoms and oomycetes. Documentation of multiple cases of domain fusions that are common to both oomycetes and diatom genomes lends additional support for the hypothesis that oomycetes and diatoms are monophyletic. Bifunctional proteins that catalyze two steps in a metabolic pathway can be used to infer the interaction of orthologous proteins that exist as separate entities in other genomes. We postulated that the novel multifunctional proteins of oomycetes could function as potential Rosetta Stones to identify interacting proteins of conserved metabolic and regulatory networks in other eukaryotic genomes. However ortholog analysis of each domain within our set of 273 multifunctional proteins against 39 sequenced bacterial and eukaryotic genomes, identified only 18 candidate Rosetta Stone proteins. Thus the majority of multifunctional proteins are not Rosetta Stones, but they may nonetheless be useful in identifying novel metabolic and regulatory networks in oomycetes. Phylogenetic analysis of all the enzymes in three pathways with one or more novel multifunctional proteins was conducted to determine the probable origins of individual enzymes. These analyses revealed multiple examples of horizontal transfer from both bacterial genomes and the photosynthetic endosymbiont in the ancestral genome of Stramenopiles. The complexity of the phylogenetic origins of these metabolic pathways and the paucity of Rosetta Stones relative to the total number of multifunctional proteins suggests that the proteome of oomycetes has few features in common with other Kingdoms.

185

16148043

The protein NONO forms heterodimers. With which proteins?

P54nrb forms a heterodimer with PSP1 that localizes to paraspeckles in an RNA-dependent manner. P54nrb is a protein implicated in multiple nuclear processes whose specific functions may correlate with its presence at different nuclear locations. Here we characterize paraspeckles, a subnuclear domain containing p54nrb and other RNA-binding proteins including PSP1, a protein with sequence similarity to p54nrb that acts as a marker for paraspeckles. We show that PSP1 interacts in vivo with a subset of the total cellular pool of p54nrb. We map the domain within PSP1 that is mediating this interaction and show it is required for the correct localization of PSP1 to paraspeckles. This interaction is necessary but not sufficient for paraspeckle targeting by PSP1, which also requires an RRM capable of RNA binding. Blocking the reinitiation of RNA Pol II transcription at the end of mitosis with DRB prevents paraspeckle formation, which recommences after removal of DRB, indicating that paraspeckle formation is dependent on RNA Polymerase II transcription. Thus paraspeckles are the sites where a subset of the total cellular pool of p54nrb is targeted in a RNA Polymerase II-dependent manner.

186

25817014

Which syndrome is associated with mutant DVL1?

Mutations in DVL1 cause an osteosclerotic form of Robinow syndrome. Robinow syndrome (RS) is a phenotypically and genetically heterogeneous condition that can be caused by mutations in genes encoding components of the non-canonical Wnt signaling pathway. In contrast, germline mutations that act to increase canonical Wnt signaling lead to distinctive osteosclerotic phenotypes. Here, we identified de novo frameshift mutations in DVL1, a mediator of both canonical and non-canonical Wnt signaling, as the cause of RS-OS, an RS subtype involving osteosclerosis, in three unrelated individuals. The mutations all delete the DVL1 C terminus and replace it, in each instance, with a novel, highly basic sequence. We showed the presence of mutant transcript in fibroblasts from one individual with RS-OS and demonstrated unimpaired protein stability with transfected GFP-tagged constructs bearing a frameshift mutation. In vitro TOPFlash assays, in apparent contradiction to the osteosclerotic phenotype, revealed that the mutant allele was less active than the wild-type allele in the canonical Wnt signaling pathway. However, when the mutant and wild-type alleles were co-expressed, canonical Wnt activity was 2-fold higher than that in the wild-type construct alone. This work establishes that DVL1 mutations cause a specific RS subtype, RS-OS, and that the osteosclerosis associated with this subtype might be the result of an interaction between the wild-type and mutant alleles and thus lead to elevated canonical Wnt signaling.

187

22057347

Are proteasome inhibitors good candidates for treatment of leukemia and solid tumors?

Inhibition of proteasome deubiquitinating activity as a new cancer therapy. Ubiquitin-tagged substrates are degraded by the 26S proteasome, which is a multisubunit complex comprising a proteolytic 20S core particle capped by 19S regulatory particles. The approval of bortezomib for the treatment of multiple myeloma validated the 20S core particle as an anticancer drug target. Here we describe the small molecule b-AP15 as a previously unidentified class of proteasome inhibitor that abrogates the deubiquitinating activity of the 19S regulatory particle. b-AP15 inhibited the activity of two 19S regulatory-particle-associated deubiquitinases, ubiquitin C-terminal hydrolase 5 (UCHL5) and ubiquitin-specific peptidase 14 (USP14), resulting in accumulation of polyubiquitin. b-AP15 induced tumor cell apoptosis that was insensitive to TP53 status and overexpression of the apoptosis inhibitor BCL2. We show that treatment with b-AP15 inhibited tumor progression in four different in vivo solid tumor models and inhibited organ infiltration in an acute myeloid leukemia model. Our results show that the deubiquitinating activity of the 19S regulatory particle is a new anticancer drug target.

188

11287619

Is there any link between CTF4 and CTF18 during sister chromatid cohesion?

Saccharomyces cerevisiae CTF18 and CTF4 are required for sister chromatid cohesion. CTF4 and CTF18 are required for high-fidelity chromosome segregation. Both exhibit genetic and physical ties to replication fork constituents. We find that absence of either CTF4 or CTF18 causes sister chromatid cohesion failure and leads to a preanaphase accumulation of cells that depends on the spindle assembly checkpoint. The physical and genetic interactions between CTF4, CTF18, and core components of replication fork complexes observed in this study and others suggest that both gene products act in association with the replication fork to facilitate sister chromatid cohesion. We find that Ctf18p, an RFC1-like protein, directly interacts with Rfc2p, Rfc3p, Rfc4p, and Rfc5p. However, Ctf18p is not a component of biochemically purified proliferating cell nuclear antigen loading RF-C, suggesting the presence of a discrete complex containing Ctf18p, Rfc2p, Rfc3p, Rfc4p, and Rfc5p. Recent identification and characterization of the budding yeast polymerase kappa, encoded by TRF4, strongly supports a hypothesis that the DNA replication machinery is required for proper sister chromatid cohesion. Analogous to the polymerase switching role of the bacterial and human RF-C complexes, we propose that budding yeast RF-C(CTF18) may be involved in a polymerase switch event that facilities sister chromatid cohesion. The requirement for CTF4 and CTF18 in robust cohesion identifies novel roles for replication accessory proteins in this process.

189

20436461

What is the Genomic Regions Enrichment of Annotations Tool (GREAT)?

GREAT improves functional interpretation of cis-regulatory regions. We developed the Genomic Regions Enrichment of Annotations Tool (GREAT) to analyze the functional significance of cis-regulatory regions identified by localized measurements of DNA binding events across an entire genome. Whereas previous methods took into account only binding proximal to genes, GREAT is able to properly incorporate distal binding sites and control for false positives using a binomial test over the input genomic regions. GREAT incorporates annotations from 20 ontologies and is available as a web application. Applying GREAT to data sets from chromatin immunoprecipitation coupled with massively parallel sequencing (ChIP-seq) of multiple transcription-associated factors, including SRF, NRSF, GABP, Stat3 and p300 in different developmental contexts, we recover many functions of these factors that are missed by existing gene-based tools, and we generate testable hypotheses. The utility of GREAT is not limited to ChIP-seq, as it could also be applied to open chromatin, localized epigenomic markers and similar functional data sets, as well as comparative genomics sets.

190

25221646

What is the target of the drug Olaparib?

Impact of DNA repair pathways on the cytotoxicity of piperlongumine in chicken DT40 cell-lines. Piperlongumine is a naturally-occurring small molecule with various biological activities. Recent studies demonstrate that piperlongumine selectively kills various types of transformed cells with minimal toxicity to non-transformed cells by inducing a high level of reactive oxygen species (ROS). ROS generates various types of DNA lesions, including base modifications and single strand breaks. In order to examine the contribution of ROS-induced DNA damage to the cytotoxicity by piperlongumine, various DNA repair-deficient chicken DT40 cell-lines with a single DNA repair gene deletion were tested for cellular sensitivity to piperlongumine. The results showed that cell lines defective in homologous recombination (HR) display hyper-sensitivity to piperlongumine, while other cell lines with a deficiency in non-homologous end joining (NHEJ), base excision repair (BER), nucleotide excision repair (NER), Fanconi anemia (FA) pathway, or translesion DNA synthesis (TLS) polymerases, show no sensitivity to piperlongumine. The results strongly implicate that double strand breaks (DSBs) generated by piperlongumine are major cytotoxic DNA lesions. Furthermore, a deletion of 53BP1 or Ku70 in the BRCA1-deficient cell line restored cellular resistance to piperlongumine. This strongly supports the idea that piperlongumine induces DSB- mediated cell death. Interestingly, piperlongumine makes the wild type DT40 cell line hypersensitive to a PARP-inhibitor, Olaparib. The results implicate that piperlongumine inhibits HR. Further analysis with cell-based HR assay and the kinetic study of Rad51 foci formation confirmed that piperlongumine suppresses HR activity. Altogether, we revealed novel mechanisms of piperlongumine-induced cytotoxicity.

191

23633213

Are thyroid hormone receptor alpha1 mutations implicated in thyroid hormone resistance syndrome?

Clinical phenotype of a new type of thyroid hormone resistance caused by a mutation of the TRα1 receptor: consequences of LT4 treatment. CONTEXT: Recently the first patients with inactivating mutations in T₃ receptor (TR)-α1 have been identified. These patients have low free T₄, low T₄, high T₃, low rT₃, and normal TSH serum levels, in combination with growth retardation, delayed bone development, and constipation. OBJECTIVE: The aim of the current study was to report the effects of levothyroxine (LT4) treatment on the clinical phenotype of 2 patients (father and daughter) with a heterozygous inactivating mutation in TRα1. SETTING AND PARTICIPANTS: Both patients were treated with LT4 for the last 5 years. To evaluate the effect of LT4 treatment, LT4 was withdrawn for 35 days and subsequently reinitiated. Data were collected from medical records, by reanalysis of serum collected over the last 6 years, and by a detailed clinical evaluation. RESULTS: Treatment with LT4 resulted in a suppression of serum TSH and normalization of serum free T₄ and rT₃, whereas T₃ levels remained elevated in both patients. In addition, there was a normalization of the dyslipidemia as well as a response in serum IGF-I, SHBG, and creatine kinase in the index patient. All these parameters returned to pretreatment values when LT4 was briefly stopped. LT4 also resulted in an improvement of certain clinical features, such as constipation and nerve conductance. However, cognitive and fine motor skill defects remained. CONCLUSION: This study reports the consequences of LT4 treatment over a prolonged period of time in 2 of the first patients with a heterozygous mutation in TRα1. LT4 therapy leads to an improvement of certain but not all features of the clinical phenotype.

192

19896475

What is the role of RhoA in bladder cancer?

The suppressive effect of Rho kinase inhibitor, Y-27632, on oncogenic Ras/RhoA induced invasion/migration of human bladder cancer TSGH cells. Urothelial cell carcinoma is the most common type of malignancy found in long-term dialysis patients and kidney transplant recipients in Taiwan. Surgical specimens of tumorous and non-tumorous bladder tissues were collected from 12 patients with bladder cancer. Increased expressions of Ras, RhoA, Akt, PI-3K were demonstrated in the tumors as compared to adjacent control tissues. To understand the impact of Ras over-expression on bladder cancer progression, human bladder cancer TSGH 8301 cells were transfected with Ras DNA. The Ras-transfected cells were then treated with either a PI-3K inhibitor (wortmannin) or Rho kinase inhibitor (Y-27632) and the expressions of Ras, PI-3K, Akt, NF-kappaB, and RhoA were analyzed. Fluorescent phalloidin staining demonstrated more intense F-actin staining in the Ras over-expressed cells than in the control cells, and the intensity of F-actin was inhibited by Y-27632. A gelatin zymography study demonstrated that the MMP-2 and MMP-9 expressions of the Ras-transfected cells were enhanced, and Y-27632 treatment reduced the levels of MMP-2 and MMP-9. Similarly, a wound healing assay revealed that the ability of cell migration was markedly increased by Ras transfection and the healing rate after treatment of Y-27632 was delayed. Our results provide evidence that Ras-induced RhoA and NF-kappaB activation was involved in the invasion/migration of bladder cancer. Through Ras and/or RhoA inhibition, there might be an opportunity for new therapeutic interventions in bladder cancer.

193

23739980

List human proteins that are subject to a dimer-to-tetramer transition.

A eukaryotic specific transmembrane segment is required for tetramerization in AMPA receptors. Most fast excitatory synaptic transmission in the nervous system is mediated by glutamate acting through ionotropic glutamate receptors (iGluRs). iGluRs (AMPA, kainate, and NMDA receptor subtypes) are tetrameric assemblies, formed as a dimer of dimers. Still, the mechanism underlying tetramerization--the necessary step for the formation of functional receptors that can be inserted into the plasma membrane--is unknown. All eukaryotic compared to prokaryotic iGluR subunits have an additional transmembrane segment, the M4 segment, which positions the physiologically critical C-terminal domain on the cytoplasmic side of the membrane. AMPA receptor (AMPAR) subunits lacking M4 do not express on the plasma membrane. Here, we show that these constructs are retained in the endoplasmic reticulum, the major cellular compartment mediating protein oligomerization. Using approaches to assay the native oligomeric state of AMPAR subunits, we find that subunits lacking M4 or containing single amino acid substitutions along an "interacting" face of the M4 helix that block surface expression no longer tetramerize in either homomeric or heteromeric assemblies. In contrast, subunit dimerization appears to be largely intact. These experiments define the M4 segment as a unique functional unit in AMPARs that is required for the critical dimer-to-tetramer transition.

194

24040872

Inhibition of which transporter is the mechanism of action of drug Canagliflozin?

Sodium-glucose co-transporter 2 inhibition in diabetes treatment: current evidence and future perspectives. Sodium-glucose co-transporters (SGLT2) are mainly expressed in the kidneys and are responsible for the renal handling of glucose load. SGLT2 inhibitors represent the latest oral agents for diabetes treatment. Their unique mechanism of action, which practically spares the insulin secretion or insulin utilization, differentiates the SGLT2 inhibitors from any existing antidiabetic agent. Thus, it is hypothesized that SGLT2 inhibitors can be effectively (and probably safely) combined with any existing antidiabetic agent (including insulin), either as monotherapy, or in dual or triple combinations. All these hypotheses are currently tested in many clinical trials. Currently dapagliflozin, one of the three most advanced SGLT2 inhibitors in the development (along with canagliflozin and empagliflozin), is already in the market in few European countries and canagliflozin has been approved from the Food and Drug Administration (FDA) in US. The evidence so far shows that SGLT2 inhibitors are equally effective to established antidiabetic agents such as metformin or sulfonylureas in their ability to lower HbA1c. On the other hand, SGLT2 inhibitors increase the possibility of genitourinary infections in type 2 diabetic individuals. Their potency in different populations and with different background therapy, but more importantly their short and long term safety remains to be seen.

195

15642542

What is the prognostic role of thyroid hormone in patients with heart failure?

Relation between free triiodothyronine/free thyroxine ratio, echocardiographic parameters and mortality in dilated cardiomyopathy. BACKGROUND: Abnormalities in thyroid function are frequent in patients with heart failure and are associated with increased mortality. However, the relation between thyroid hormone levels and echocardiographic parameters has not been investigated sufficiently. AIM: The aims of this study were to investigate the correlations of thyroid hormone levels with echocardiographic parameters and to evaluate their associations with subsequent mortality in a group of patients with dilated cardiomyopathy (DCMP). METHODS: Serum levels of thyroid hormones were measured in 111 consecutive patients with DCMP (35 female, 76 male, mean age: 62+/-12 years). All patients underwent echocardiographic examination and were followed-up for a period of 12+/-8 months. RESULTS: Twenty-three patients (21%) had abnormalities in thyroid function tests. Free triiodothyronine (fT3)/free thyroxine (fT4) ratio was significantly correlated with most of echocardiographic parameters, such as chamber diameters and ejection fraction. Sixteen patients (14%) died during the follow-up period; their fT3/fT4 ratio was significantly lower than the patients who survived (1.31+/-0.37 vs. 2.01+/-0.72, p<0.001). A fT3/fT4 ratio of <or=1.7 was associated with an increased risk of mortality (p<0.001), independent of other prognostic markers. Sensitivity, specificity, positive and negative predictivity of fT3/fT4 ratio <or=1.7 for cardiac mortality were 100%, 71%, 36% and 100%, respectively. CONCLUSION: Determination of FT3/FT4 ratio may be a valuable and simple predictor for identification of patients with DCMP who are at high risk of subsequent mortality.

196

17655857

which mutations of phospholamban gene have been found to cause hypertrophic cardiomyopathy?

Genetic screening of calcium regulation genes in familial hypertrophic cardiomyopathy. Genes encoding Ca(2+) regulatory proteins responsible for Ca(2+) homeostasis have been suggested as possible candidates for FHC. Mutations in sarcomere genes account for approximately 50% of all FHC cases indicating other genes, including those involved in Ca(2+) handling, may account for the remainder. The aim of this study was to identify causative mutations in genes involved in Ca(2+) regulation in patients with familial hypertrophic cardiomyopathy (FHC). An Australian cohort of 252 unrelated familial hypertrophic cardiomyopathy patients were screened for mutations in the Ca(2+) regulatory genes, sorcin (SRI), calstabin (FKBP1B), calsequestrin (CASQ2), phospholamban (PLN), sarcolipin (SLN), calreticulin (CALR3) and calmodulin (CALM). A total of 17 exonic DNA variants were identified in the 7 Ca(2+) regulatory genes studied, of which 4 were considered of pathogenic significance. Two novel mutations in the CALR3 gene were identified (Lys82Arg, Arg73Gln) and one truncation mutation in the PLN gene (Leu39Ter). A variant was also identified in the CASQ2 gene (Asp63Glu). These four variants were all novel, resulted in changes in conserved amino acids and were not identified in a normal population. In conclusion, mutations in Ca(2+) handling genes are an infrequent but important cause of FHC. DNA variants in Ca(2+) genes may also be involved as modifying factors in phenotype development. Further evaluation of the role of defects in Ca(2+) regulation will shed light on the molecular pathogenesis of FHC.

197

18817898

Which gene strand is targeted by transcription-coupled repair (TCR)?

Yeast Elc1 plays an important role in global genomic repair but not in transcription coupled repair. Transcription coupled repair (TCR) is a nucleotide excision repair (NER) pathway that is dedicated to repair in the transcribed strand of an active gene. The genome overall NER is called global genomic repair (GGR). Elc1, the yeast homolog of the mammalian elongation factor elongin C, has been shown to be a component of a ubiquitin ligase complex that contains Rad7 and Rad16, two factors that are specifically required for GGR. Elc1 has also been suggested to be present in another ubiquitin ligase complex that lacks Rad7 and Rad16 and is involved in UV-induced ubiquitylation and subsequent degradation of RNA polymerase II. Here we show that elc1 deletion increases UV sensitivity of TCR-deficient cells but does not affect the UV sensitivity of otherwise wild type and GGR-deficient cells. Cells deleted for elc1 show normal NER in the transcribed strand of an active gene but have no detectable NER in the non-transcribed strand. Elc1 does not affect UV-induced mutagenesis when TCR is operative, but plays an important role in preventing the mutagenesis if TCR is defective. Furthermore, the levels of Rad7 and Rad16 proteins are not significantly decreased in elc1 cells, and overexpression of Rad7 and Rad16 individually or simultaneously in elc1 cells does not restore repair in the non-transcribed strand of an active gene. Our results suggest that Elc1 has no function in TCR but plays an important role in GGR. Furthermore, the role of Elc1 in GGR may not be subsidiary to that of Rad7 and Rad16.

198

10754001

Abnormalities in which chromosomes were linked to the Moyamoya disease?

Linkage of familial moyamoya disease (spontaneous occlusion of the circle of Willis) to chromosome 17q25. BACKGROUND AND PURPOSE: Moyamoya disease is a cerebrovascular disease of unknown cause that mainly affects Japanese children. The incidence of familial occurrence accounts for 9% of cases. The characteristic lesions of moyamoya disease are occasionally seen in neurofibromatosis type 1, of which the causative gene (NF1) has been assigned to chromosome 17q11.2. METHODS: To determine whether a gene related to moyamoya disease is located on chromosome 17, we conducted microsatellite linkage analyses on 24 families containing 56 patients with moyamoya disease. Leukocyte DNA extracted from the family members was subjected to polymerase chain reaction for a total of 22 microsatellite markers on chromosome 17. The amplified polymerase chain reaction fragments were analyzed with GeneScan on an automated sequencer. RESULTS: Two-point linkage analysis gave a maximum log(10) odds (LOD) score of 3.11 at the recombination fraction of 0.00 for the marker at locus D17S939. The affected pedigree member method also showed a significantly low P value (<1. 0x10(-5)) for the 5 adjacent markers at 17q25. Multipoint linkage analysis also indicated that the disease gene is contained within the 9-cM region of D17S785 to D17S836, with a maximum LOD score of 4. 58. CONCLUSIONS: A gene for familial moyamoya disease is located on chromosome 17q25.

199

18824513

Which is the branch site consensus sequence in U12-dependent introns?

Mutational analysis of the U12-dependent branch site consensus sequence. Highly conserved sequences at the 5' splice site and branch site of U12-dependent introns are important determinants for splicing by U12-dependent spliceosomes. This study investigates the in vivo splicing phenotypes of mutations in the branch site consensus sequence of the U12-dependent intron F from a human NOL1 (P120) minigene. Intron F contains a fully consensus branch site sequence (UUCCUUAAC). Mutations at each position were analyzed for their effects on U12-dependent splicing in vivo. Mutations at most positions resulted in a significant reduction of correct U12-dependent splicing. Defects observed included increased unspliced RNA levels, the activation of cryptic U2-dependent 5' and 3' splice sites, and the activation of cryptic U12-dependent branch/3' splice sites. A strong correlation was observed between the predicted thermodynamic stability of the branch site: U12 snRNA interaction and correct U12-dependent splicing. The lack of a polypyrimidine tract between the branch site and 3' splice site of U12-dependent introns and the observed reliance on base-pairing interactions for correct U12-dependent splicing emphasize the importance of RNA/RNA interactions during U12-dependent intron recognition and proper splice site selection.