test_extracted_captions/chen2023novo.json

{"CAPTION FIG6.png": "'Figure 6: Colled-coil interfaces are suitable to withstand mechanical stress from orthogonal directions (A) A model of how 3-helix bundles would be able to bear mechanical stress differently compared with the microtubules. The bending curvatures and gaps are exaggerated forustration purposes.\\n\\n(B) A schematic of wild-type and mutant sperm doublet structures highlighting the Takitin 5 bundles and the partial redundancy.\\n\\n'", "CAPTION FIGS1.png": "'Figure S1. Workflow of data processing, related to Figures 1, 2, and 3 and STAR Methods (A) Tift series composed of 25 2D projections were recorded [scale bar: 100 nm]. The image shows the midpiece of sperm flagella that contains mitochondria around the axoneme. Gold beads on the tilt images are indicated (green arrowhead). The fiducial gold beads were used to align the tilt images. (B) 3D micrograms were reconstructed, and subroutines were picked along the microtubules (scale bar: 100 nm). (C) 3D elastifica and refinement were performed using and average the subutemogram for the 96 nm-propagating structures of the mouse sperm doublets. Four views of the 96 nm-repeating structure of doublets from EHNA-treated sperm are shown for the 3D reconstruction generated using RELION3 as reported previously.10 Gold-standard Fourier shaft correlation (FSC) curve calculated between half maps of mouse sperm doublets is shown. The resolution was estimated as 26 A (FSC = 0.143). (D) Two slices of the 96 nm-repeating structure of doublets looking along and perpendicular to the filament axis. Note the red line in the top panel indicates the plane of the bottom slice, and periodic structures are observed inside the microtubules. The coordinates were recartered on the 48-nm repeats and imported into RELION4. In the top panel, note that the features further away from the microtubules are blunier, suggesting that there are conformational heterogenities and that they are resolved at lower resolutions. (E) The initial father3D job of the 48 nm-repeating structures was performed using RELION4.21\\n\\n(F) The 3D reconstructions were matched to the 2D projections of individual particles in the raw tilt images, and this step refined both the geometric and optical parameters of the tilt series (-H). (G) Another round of sub-tetromogram averaging was performed based on refined tilt series. No additional improvement was observed after 3 rounds of refinement and Refine3D as shown in (F)-(G).\\n\\n'", "CAPTION FIGS4.png": "'Figure S4.\u2014 Rigid-body fitting of 29 identified MIPs from bovine trachea cilia into the density map of mouse sperm doublet, related to Figures 2 and 3 and STAR Methods (A-F) Models of 28 known MIPs from bovine trachea cilia (PDB: 7MRC)\\\\({}^{13}\\\\) are fitted into the density map of mouse sperm doublet. The viewing angles for all panels are shown. For proteins that have multiple \u03b1-helices (CFAP161, RIBC2, CFAP53, MNS1, CFAP21, NME7, CFAP141, EFHC1, EFHC2, ENKUR, CFAP210, EFCABE, GFAP45, PACRB, and TEKIN 1\u20134), the arrangement of secondary structures matches densities in sperm doublets. The overall shapes of \u03b2-sheets in \u03b2-sheet-rich proteins (CFAPS2 and CFAP20) match the densities, and these proteins are highly conserved in axoremes. For the proteins that contain random coils, we did observe matching features in the maps and were able to trace some of the main chairs at the current resolution (CFAP96, SPAG8, CFAP107, FAM166B, Pierce1, Pierce2, CFAP128, CFAP276, and TEKIFIP1).\\n\\n'", "CAPTION FIGS5.png": "'Figure S5: Characterization of the 16 nm-repeating structures of doublets from mouse sperm, related to Figure 2 (A and B) Gold-standard Fourier shell correlation (FSC) curves were calculated using half maps of 16 nm-repeating structures of A-bLuMe and B-bLuMe. The resolution was estimated as 6.0 and 7.7 \\\\(\\\\AA\\\\), respectively (FSC = 0.143). The Nyquist limit is 5.26 \\\\(\\\\AA\\\\).\\n\\n(C-E) The local resolution map was calculated from the two half maps of 16 nm-repeating structures of A-bLuMe using RELION4. The viewing angles for (G) and (E) are shown in (C) (black arrow). These viewing angles are similar to Figures 1A, 1D, and 1E, respectively.\\n\\n(F-H) The local resolution map was calculated using the two half maps of 16 nm-repeating structures of B-bLuMe using RELION4. The viewing angles for (G) and (G) are shown in (F) (black arrow). The viewing angles of (F) and (G) are similar to Figures 1A and 1F, respectively.\\n\\n'", "CAPTION FIGS6.png": "'Figure S6.: Tektin 5 and CCDC105 likely form sperm-specific 3-helix bundles associated with the A-tubule, related to Figures 2 and 3 (A) After unbiased matching, Tektin 5 was scored as the 45 h of the predicted structures out of 21,615 proteins from the mouse proteome, marked by cross-correlation scores (the top 10 are shown). Tektin 1-4 were ranked at 47-10 due to their similar tertiary structures.\\n\\n(B) Typical elastic positions (#1-4 and e/b) from the same search. Usually, these are proteins with long single helices that do not match the gaps observed in the map. Also, they do not explain the 3-helix bundles. The fitting of Tektin 5 into the same densities is shown for comparison.\\n\\n(C) The structure of CCDC105 directly predicted by AlphaFold2 (left) is compared with the predicted complex formed by two CCDC105 molecules (right). The half-length CCDC105 molecule in the complex is colored based on the per-residue confidence scores (predicted local distance difference test (pLDDT), red: high confidence) from the AlphaFold2 prediction. The three P-loops have medium confidence scores (green), suggesting the exact conformations of these loops may not be accurately predicted. However, the presence of these structured loops is conclusively confident based on the conserved proline residues (see the sequence alignment in D) and the matching protrusion densities observed in our maps (Figure 2G). Note that the conformations of the three proline-rich loops differ in these two predictions. These differences could be caused by the presence of neighboring molecules during AlphaFold2 analyses.[7]\\n\\n(D) The sequence alignment of CCDC105 from the mammals (H. sapiens, _M. musculus, B. taurus, S. scrota, and F. catus), zebrafish (D. rerio), and sea urchins (S. roupurakis). The three proline-rich loops are marked above the sequences.\\n\\n(E) The models of CCDC105 and Tektin 5 are fitted into the densities of the 3-helix bundle at their ribbon, where the former model explains the extra protrusions and orientation/lengths of helices of the densities, but the latter does not.\\n\\n'", "CAPTION FIGS8.png": "'Figure S8: Biochemical extractions of proteins from mouse sperm, related to Figure 3 (A) SDS-PAGE analyses of protein extractions from mouse sperm using 0.1% Triton in PBS (E1), 0.6 M NaCl in PBS (E2), 0.6 M KClN in PBS (E3), 8 M urea (E4), and 10% SDS (E5).\\n\\n(B) Western blot analyses of protein extractions from mouse sperm using an antibody against a-tubules. Note that strong bands were detected only in E3 and E4, suggesting the microtubule structures were stable in Triton and high NaCl buffers and dissonhol completely in KClN/urea solutions.\\n\\n(C) Bar chart of the number of proteins identified by MS (protein control in each fraction (E1-ES) and biological replicate. We identified a total of 1,677 mouse proteins, with a range of 772 to 1,326 proteins identified in each individual fraction and replicate.\\n\\n(D) Hathamp of proteins with significant changes between any two fractions (absolute log2 fold change [log2-FC] > 1, adjusted p value < 0.05), listed by fractions (E1-ES) and biological replicate, and clustered by correlation of intensity profile. Proteins are colored by the log2,FC in protein intensity normalized to the row median (red, increased intensity; blue, decreased intensity; gray, not detected). Cluster identification _narrows_ (cluster ID) are labeled (err).\\n\\n(E) Hathamp of gene ontology (GC) enrichments among the significantly changing proteins identified in each cluster from (D) that to right: cluster ID 1\u20136, as labeled in ID. GO terms were curated from the top 4 enrichment terms per cluster, and non-redundant terms were selected by an automated clustering procedure (see STAR Methods). Increased shading reflects increased significance of the enrichment term. The number of proteins per enrichment term is shown in white IR significant (adjusted p value < 0.05) and gray if not significant (adjusted p value > 0.05). A bar chart plotting the number of total genes in each cluster ID is included.18\\n\\n(F) Log2 protein intensities (y axis) for eight mouse proteins as quantified by MS in each fraction (E1\u2013ES) and biological replicate (colored dots; maximum n = 3).\\n\\n'", "CAPTION FIGS2.png": "'Figure S2: Characterization of the 48 nm-repeating structure of doublets from mouse sperm, related to Figures 1, 2, and 3\\n\\n(A) Gold-standard Fourier shell correlation (FSC) curves were calculated between half maps of mouse sperm doublets. The resolutions were reported as FSC = 0.148. Note the FSC curves resulting from the iterative frame alignment and CTF refinement between the second and third Refrea3D [abs were not shown for the activity of the figure. Further refinement after the third Refrea3D did not improve the resolution or the quality of the map.\\n\\n(B) The local resolution map of mouse sperm doublets was calculated by RELLOU4. The ribbon region has the highest resolutions. Densities in the A-tubule have higher resolutions than the ones from the B-tubule.\\n\\n(C) Equivalent longitudinal cross-section views of doublets from mouse sperm and bovine trachea cilia (EMDE: EMD-24664) are shown. [(ii)] The latter was low-pass filtered to 7.5 \u00c5, and comparable details of the secondary and tertiary structures of the MPAs are observed.\\n\\n(D) The reconstruction of mouse sperm doublet (gray) is overlaid with the bovine trachea doublets (yellow). The mouse sperm-specific densities are highlighted (dashed axis). The broken helical bundles and the curved helical bundles inside the A-tubule of mouse sperm doublets along the microtubule axis are shown. The discontinuous parts of the broken helical bundles are indicated (dashed rectangle). Note that the curved bundles have one straight and two curved groups of densities in every 48-nm repeat (outlined using dashed shapes).\\n\\n'", "CAPTION FIG1.png": "'Figure 1: The 3D reconstructions of mouse and human sperm doublets revealed novel MIPs (A and B) Transverse cross-section views of the doublets of mouse (A) and human (B) sperm. Conserved sperm MIP densities are highlighted (pink, blue, and green), and the corresponding viewing angles of (C)\u2013(E) are indicated (colored arrowheads). The 3-helix densities in A-tubule shared with bovine trachea doublets (EMD-24664) are colored (yellow).\\\\({}^{13}\\\\) Divergent sperm densities are also indicated (red dashed shapes). Individual protofilaments of the doublets are labeled as A1\u2013A13 and B1\u2013B10. (C\u2013E) Zoom-in views of the conserved sperm MIP densities along the longitudinal axis. In (C), mouse sperm-specific densities are indicated and labeled (red dashed shapes; see more in Figures S2 and S3). In (E), although the situations are 8 nm apart from one another, the overall periodicity is 48 nm. See also Figures S1, S2, and S3.\\n\\n'", "CAPTION FIGS3.png": "'Figure S3. Characterization of the 48 nm-repeating structure of doublets from human sperm, related to Figure 1 (a). The gold-standard Fourier shell correlation (PSC) curve was calculated between half maps of mouse sperm doublets. The resolution was estimated as 10.3 A (FSC = 0.143).\\n\\n(B) The local resolution map of human sperm doublets was calculated by RELION4. The ribbon region has the highest resolutions. Densities in the A-tubule have higher resolutions than the ones from the B-tubule.\\n\\n(C) Equivalent views of doublets from human sperm and bovine trachea cilia (EMOE: EMD-24664) are shown. [13] The latter was low-pass filtered to 10 A, and comparable details of the secondary and tertiary structures of the MMPs are observed.\\n\\n(D) The reconstruction of human sperm doublet (blue) is overlaid with the bovine trachea doublets (yellow) at low and high thresholds.\\n\\n(E) The two broken bundles inside the A-tubule in human sperm are shown at a low threshold (see the corresponding mouse densities in Figure S2D).\\n\\n(F) The curved helical bundles contain one straight and two curved groups of densities inside the A-tubule of human sperm. Human sperm-specific densities were observed to connect one curved bundle to the lumen of A-tubule.\\n\\n(G) The human sperm doublets overlaid with mouse sperm doublets are shown. The inconsistent densities are outlined (dashed line) (also see Figures S2D and S2F).\\n\\n'", "CAPTION FIGS7.png": "'Figure S7. DUSP proteins in the A-tubule, related to Figure 3 (A) at a lower threshold compared with Figure 3C, densities connecting the N-terminal residues of the slanted Takin Ss (magenta models) and the DUSPs (blue models) are observed.\\n\\n(B) The DUSPs is fitted into the globular domain, and three orthogonal views are shown. Other homologous DUSP proteins fit well into the density because of similar tertiary structures [DUSP 3, 13, 14, 18, 21, and 29].\\n\\n'", "CAPTION FIG2.png": "'Figure 2: _De novo protein identification of sperm MIPs assisted by AlphaFold2_\\n\\n(A) Conserved densities in mouse and human sperm were segmented from the averages of 16-nm repeats of mouse sperm doublets and searched in the AlphaFold2 library of the mouse proteasome (21,615 proteins).\\n\\n(B) The predicted structure of Takith 5 based on AlphaFold2 was fitted into the continuous 3-helix bundle.\\n\\n(C) Modeling of a complex formed by a full-length Takith 5 and a truncated one (N-Tait 5: aa 1-149) using CableFold.23\\n\\n(D) Fitting and modeling of Takith 56 into the 3-helix bundle identifies in the A-t-'", "CAPTION FIG5.png": "'Figure 6: Characterization of mutant TeMFS -/- sperm\\n\\n(**A**) The percentages of motile sperm from wild-type and TeMFS knockout mice (>200 cells were counted for each mouse, three knockout -/- mice and two wild-type mice were analyzed, and the pool percentage and 95% confidence intervals Wilson/Brown method were shown).\\n\\n(**B**) The percentages of bent sperm from wild-type and TeMFS knockout mice (>200 cells were counted for each mouse, three knockout -/- mice and two wild-type mice were analyzed, and the pool percentage and 95% confidence intervals by Wilson/Brown method were shown). Two examples of bent sperm are shown.\\n\\n(**C**) An overlay of wild-type models with the densities of TeMFS -/- sperm around the slanted bundles. The continuous 3-helix bundle assigned as TeMFS (high occupancies) and slanted helical bundles (low occupancies) are shown. The densities corresponding to the DUSP proteins are barely resolved. Note that there are substantially less densities for these models compared with Figure 3C.\\n\\n(**D**) An overlay of wild-type models with the densities of TeMFS -/- sperm around the curved bundles. The occupancies of the curved bundles are lower than the other MIPs and tubulins. Note that there are substantially less densities for these models compared with Figure 3D.\\n\\n(**E**) The two broken 3-helix bundles have lower occupancies compared with the surrounding MIPs and tubulins. Note that there are substantially less densities for these models compared with Figure 3A.\\n\\n'", "CAPTION TABNA.png": "'\\n\\n**Acknowledgments**'", "CAPTION FIG4.png": "'Figure 4. Sperm doublet are compared at microtubules and extensive cact-cell bundles\\n\\nFig 5. The pus and rhinus arcade of Takin 5 were named in a fixed C terminal of the protein.\\n\\n[MISSING_PAGE_POST]\\n\\n'", "CAPTION FIG3.png": "'Figure 3.: **Conformational plasticities of Tektin 5**(A) The two broken 3-helix bundles could be explained by two complete and a third partial copies of Tektin 5 (dashed rectangles) per 48-nm reagent, instead of three Tektin in the continuous 3-helix bundle.\\n\\n(B) The AlphaFold2 medial of mouse Taktin 5 was fitted into the slanted helical damilies. Sequence alignment of Tektin 5 from _M. musculus_, _H. sapiens_, _B. fauvus_, and _F. cada_ is shown from G133 to F151 (the numbering of ambe acids is based on _M. musculus_ Tektin 5). The conserved Gly137, Gly143, and Gly150 are near the turning point of the bent \u03b1-helix.\\n\\n(C) The fitting of Tektin 5 and DUSF3 protein (its homologs are also possible candidates) into the 16 nm-repeating features; see the same view of the map in Figure 1C.\\n\\n(D) Three modified Taktin 5 were fitted into the densities of the curved bundle in the mouse sperm doublet (as indicated in Figure 1A). The intact intermolecular interaction strength reference, N termini of the Taktin 5s, and curved 2-helix segments are indicated (arrows). Nearby MIPs shared between mouse sperm flagella and bovine trachea cilia are also colored and labeled [MMET, CFAP161, and SPAGB].\\n\\n(E) The cross-section schematic is shown. The highlighted models of (A)\u2013(D) are indicated using arrows. See also Figures 51, 52, 54, 56, 57, and 58.\\n\\n'"}