EMBO/sd-character-level-ner huggingface dataset:
Figure 5Joint analysis of WT and kit mutant MC. A, tSNE plot colored by WT and kit mutant MC subpopulations. Representative GO terms of genes upregulated in MC1 (WT, red bars) or KMC1 (kit mutant, blue bars) from the uninjured hearts, respectively. Dotted lines indicate P= 0.05. C, Bar plots showing proportion of each WT and kit mutant MC subpopulation in total nonCM at each time point. D, F, Violin plots showing the expression of il1b in MC1 (D), and ctsd expression in MC (F). E, G, Expression of il1b (E) and ctsd (G) in WT and kit mutant hearts determined by qRT-PCR. Joint analysis of WT and kit mutant FB. H, tSNE plot colored by WT and kit mutant FB subpopulations. I, Bar plots showing proportion of each WT and kit mutant FB subpopulation in total nonCM at each time point. Violin plots showing expression of col12a1a in FB3 (J) Expression of col12a1a (K) in WT and kit mutant hearts determined by qRT-PCR. mif expression in FB (L). Expression of mif (M) in WT and kit mutant hearts determined by qRT-PCR. N, Bar plots showing relative proportion of each WT and kit mutant eEC subpopulation in eEC at each time point. O, Violin plots showing expression of junba in eEC. P, Expression of junba in WT and kit mutant hearts determined by qRT-PCR. Q, Representative image of hearts from 7 dpi tcf21:nucGFP transgenic fish treated by PBS liposome, Clodronate liposome and Dex, respectively. The red boxes mark the peripheral area, and the white boxes mark the injury area and are shown in their respective zoom-in images to the right.. White dashed lines indicate approximate resection plane. R, Quantification of the number of tcf21:nucGFP positive cells in the boxed areas in U. N = 5 hearts.
Phosphorimage analysis of in vitro CK2 kinase assays with protein substrates.figf5Phosphorimage analysis of in vitro CK2 kinase assays with protein substrates. All reactions were performed at 30 °C. Numbers indicate the concentration (nM) of product for each kinase reaction. (a) AHCYL2 substrate (25 ng μl−1) with 100 μM ATP and 10 nM CK2α ± 11 nM CK2β for 6 min. (b) NAP1L3 substrate (15 ng μl−1) with 100 μM ATP and 10 nM CK2α ± 11 nM CK2β for 6 min. (c) NIPBL substrate (50 ng μl−1) with 100 μM ATP and 5 nM CK2α ± 11 nM Pin1 for 5 min. Full gels corresponding to those in panels a-c are in Supplementary Figures 19 and 25.
Figure 7(A) NSC-34 and SH-SY5Y cells were infected with EV-A71 S41 at MOI 30 and 0.01, respectively. At 48h.p.i., the cells were fixed and permeabilized prior to immunostaining. B) Proximity ligation assay. NSC-34 and SH-SY5Y cells were transfected with LucEV-A71 replicon. Cells were fixed and permeabilized at 24h.p.t. prior to PLA staining using antibodies specific to Rac1, 3C and 3D. (C) Total cell lysates were pulled down with anti-Rac1 or IgG isotype control antibody prior to immunoblotting. Uninfected cells serve as negative control. IP: immunoprecipitation; IB: immunoblotting.
The morphology of the UIS4-positive LS‐TVN.Figure 2Hepatoma cells infected with Plasmodium berghei ANKA parasites were fixed and stained with anti-UIS4 and anti-HSP70 A) Some UIS4‐positive vesicles are tethered to the PVM as 'beads‐ona‐string'. Infected cell fixed 24 h post‐infection. B) The UIS4‐mCherry features we detect in live imaging of UIS4‐mCherry‐expressing P. berghei, i.e. elongated membrane clusters (left), vesicles (center) and tubules (right) are also apparent in fixed wildtype parasites stained with anti‐UIS4 antibody. Infected cells 24 h post‐infection. Scale bars, 10 µm. Hepatoma cells infected with Plasmodium berghei ANKA parasites were fixed and stained with anti-UIS4. C) Often the LS‐TVN is a composite of vesicles, tubules and membrane clusters. Infected cell 22 h post‐infection. D) The LS‐TVN detectable in primary hepatocytes infected with UIS4‐mCherry‐expressing P. berghei also consists of elongated membrane clusters (yellow arrowhead), vesicles (white arrow) and tubules (yellow arrow). Infected cell 18 h post‐infection. E) Dynamic tubules (yellow arrow) are detectable with UIS4‐mCherry in infected primary hepatocytes 24 h post‐infection. Location of parasite is marked with a yellow asterisk. Scale bars, 10 µm.
Fig.Figure 3Littermate control and δWD mice (n = 5) were challenged with IAV X31 (103 pfu). (A) At the indicated time points, cytokine mRNA transcripts in lung tissue (n = 5) were evaluated by qPCR. Littermate control and δWD mice (n = 5) were challenged with IAV X31 (103 pfu). (B) Representative lung sections from animals (n = 6) taken at 3 d.p.i. were stained by IH for neutrophils (Ly6G) or neutrophil extracellular traps (NET; anti-H3). Sections at 7 d.p.i. were stained for macrophages (Iba-1). Littermate control and δWD mice (n = 5) were challenged with IAV X31 (103 pfu). C) BAL (n = 5) was taken at 2 d.p.i and evaluated by flow cytometry, with pre-gating on CD45+. The percentage of neutrophils (CD11b+, Ly6G+) cells is shown ±SEM and were compared using Mann-Whitney U test (*P<0.05). Littermate control and δWD mice (n = 5) were challenged with IAV X31 (103 pfu). (D) Single cell suspensions were prepared from lungs taken at 5 d.p.i and evaluated by flow cytometry, with pre-gating on CD45+. The percentage of macrophage/monocytes (CD11b+, F4/80+) cells is shown ±SEM and were compared using Mann-Whitney U test (*P<0.05). (E) δWD mice were treated with either anti-PCDA-1 (to deplete plasmacytoid DC) or an isotype-matched control. Litter-mate control mice were used as comparator. Weight loss was measured at the indicated days p.i. (n = 5).
Figure 5(A) The Hrr25:Mam1 complex with Mam1 shown as orange ribbons, and Hrr25 shown as a white surface with Mam1-binding interface #1 shown in green and #2 in magenta. (B) Stereo view of interface #1, which involves hydrophobic interactions by Mam1 L92, L97, L100, and I104, and a hydrogen-bond network between Mam1 Q101/E108/R131 and Hrr25 residues 12-15. (C) Stereo view of interface #2. Hrr25 residues H25 and E34 participate in a buried hydrogen-bond network with Mam1 residues R149 and Y158, and are surrounded by hydrophobic residues on both proteins. For clarity, Mam1 helix 4, on which R149, Y153, and Y158 are located, is not shown. (D) Pulldown assay with in vitro-translated Mam187-191 (wild-type and alanine mutants) and Hrr251-394 K38R. Mam1 mutations also disrupt Hrr25 binding when the two are co-expressed in E. coli (Figure EV6B). Bottom: Sporeviability of MAM1 mutant strains (see Table EV2 for strains; strains tested are KC549, KC560, KC566, KC552, KC554, KC556, and KC558; 46-48 tetrads were dissected for each strain). "--" indicates that this mutation was not tested for sporeviability.
Figure 6(A) B6.K18-hACE2IP-THV mice were immunized with LV::S via i.m. or i.n. at wk 0 and boosted via i.m. or i.n. at wk 5 (n = 5/group) with non-integrative LV::S. Control mice were injected i.m. i.n. with an empty LV (sham). Viral RNA contents were determined by conventional E-specific RT-PCR at 3 dpi, in the brain, lung and nasal washes. Data information: Statistical significance was evaluated by Mann-Whitney test (*= p < 0.05, **= p <0.01, ****= p < 0.0001, ns = not significant). (B) Lung histology in B6.K18-hACE2IP-THV mice, vaccinated with LV::S after SARS-CoV-2 inoculation. H&E (rows 1 and 3) (scale bar: 500 μm) and NCoV-2-specific IHC (rows 2 and 4) (scale bar: 50 μm) staining of whole lung sections (scale bar: 50 μm) from the primed (i.m.), boosted (i.n.) and challenged B6.K18-hACE2IP-THV mice compared with their sham controls. H&E and NCoV-2-specific IHC were performed on contiguous sections. The IHC fields correspond to the rectangles in the corresponding H&E images above them. A representative NCoV-2-specific IHC on a lung section from a non-infected (NI) mouse is also shown. (C) Heatmap representing the histological scores for various parameters in LV::S-vaccinated or sham mice at 3 dpi (n = 6/group).
Figure 5A. Domain organization of human FAM111B and overview of disease-associated heterozygous missense mutations. B. U2OS/GFP-FAM111A WT cells treated or not with DOX were subjected to GFP IP followed by immunoblotting with indicated antibodies. C. As in (B), using U2OS/GFP-FAM111B WT cells. D. Purified recombinant FLAG-FAM111A and FLAG-FAM111B proteins were incubated separately or mixed, subjected to IP with FAM111A antibody and immunoblotted with FLAG antibody. E. Immunoblot analysis of U2OS/GFP-FAM111B cell lines treated or not with DOX in the absence or presence of Z-VAD-FMK. F. Cells in (E) were pulse-labeled with EdU, stained with DAPI and analyzed for DAPI and EdU signal intensity using QIBC. Cells analyzed using QIBC. G. Quantification of data in (F) for S phase (EdU-positive) cells (red bars, mean (A.U., arbitrary units); n>2000 cells per condition). Cells stained with DAPI and analyzed for DAPI signal intensity using QIBC. H. cells were pulse-labeled with EU. Cells analyzed using QIBC. I. Quantification of EU incorporation in cells in (H) (red bars, mean; n>2000 cells per condition). J. U2OS/GFP-FAM111B cell lines (Figure 5E) were fixed at the indicated times after DOX treatment and stained with crystal violet. K. Cells in (E) were fixed, immunostained with γ-H2AX antibody and subjected to QIBC analysis of γ-H2AX signal intensity (red bars, mean; n>2000 cells per condition). L. Immunoblot analysis of purified recombinant FLAG-FAM111B proteins and their auto-cleavage products.
Figure 3 (A) Western blot of the SUMO conjugation reaction in the presence of different oligonucleotides using the minimal Smc5/ARM-Nse2 construct. Reactions were run at 30°C and stopped after 60 minutes by adding SDS-loading buffer. 25b (μM), 34b (μM) and 50b (μM), stands for a 25, 34 and 50 bases oligonucleotides, respectively, and the indicated concentration is in μM units. 5kb (nM), stands for the virion ϕx174 and the indicated concentration is in nM units (N-S1, cNse4-SUMO1; N-2S1, cNse4-2SUMO1; and pS1, poly-SUMO1) (*Overexposed chemiluminescent signal)  (B) Left, Ubc9-thioester formation in the presence of E1, E2 enzymes, Alexa488-SUMO1 and ATP. Right, single turnover reaction of the SUMO conjugation reaction in the presence or absence of ssDNA (50b) using Arm/Smc5-Nse2 as E3. Samples were run in the presence (below) or absence of β-mercaptoethanol (above)  (C) Ribbon representation of the complex between the ARM domain of Smc5 (yellow and orange) and Nse2 (pink) (PDB 3HTK) (Duan et al, 2009). Lysine residues forming the positive-charged patch in the surface of the coiled-coil Smc5 ARM are labeled and shown in stick representation (blue). Zinc atom in the Nse2 RING domain is depicted as a yellow sphere  (D) Bar diagram representation of the SUMO conjugation rates of activity assays of Arm/Smc5-Nse2 KE mutants in the presence (orange bars) or absence (red bars) of ssDNA (virion ϕx174), relative to wild-type (set to 1). Reactions rates were performed at least in three different independent experiments. Data values are mean ± s.e.m. and n=3 technical replicates. Significance was measured by a two-tailed unpaired t-test relative to wild-type. *P<0.05, **P<0.01, ***P<0.001  (E) DNA binding properties of wild-type, K337E/K344E/K764E and K743E/K745E Arm/Smc5-Nse2 mutants, were determined by electrophoretic mobility shift assays (EMSA) saturation experiments. Protein complexes were incubated for 30 minutes at 30°C before loading the agarose gel electrophoresis. Numbers above gel indicate the molar ratio (x103) of protein over ssDNA (virion ϕx174) in each lane
Figure 5A. LC3C specific sensor mCh-AS3_67 is recruited to Salmonella (SFH2) escaping SCV. Salmonella SFH2 strain is carrying a GFP-reporter gene activated by availability of glucose-6-phosphate in the host's cytosol. B. Salmonella with prolonged cytosolic residence and strong activation of the GFP-reporter gene (green arrow) are not recognized by the LC3C specific sensor mCh-AS3_67. C. A subpopulation of hyper proliferative Salmonella in the cytosol is coated with endogenous LC3C. In contrast, endogenous LC3B is enriched in spots in close proximity to the cytosolic bacteria. D. Sensitivity of LC3C specific sensor mCh-AS3_67 is increased upon fusion of membrane-tethering FYVE domain or PB1 oligomerization domain. E. Introduction of the PB1 domain to mCh-AS3_p62 increases its performance in recognizing endogenous LC3B. F. High-performance sensor mCh-PB1-AS3_67 was transiently expressed in Salmonella infected HeLa cells to monitor xenophagy events.
Molecular interactions of the human PCI-MPN core of eIF3 in the 43S PIC.Figure 5A Isolated map and molecular model of the eIF3 PCI-MPN core color coded as in Fig 2. Structural hallmarks are indicated, and a scheme shows the composition of the lobes. B, C Interactions of eIF3a, eIF3c and eIF3d with the ribosome: (B) shows an overview of the structure and zoomed views highlighting the interactions of eIF3a, eIF3c, the eIF3d N-terminal tail and the eIF3d cap-binding domain with the 40S, (C) shows molecular details of eIF3a interacting with eS1; eIF3c interacting with rRNA h22 and eIF3c and the N-terminal tail of eIF3d with the Zn-knuckle domain of eS27. D Interactions of the eIF3d N-terminal tail with the PCI-MPN core.
GRK2 is essential for skeletal growth plate development.Figure 3(A) GRK2 immunohistochemistry in a control 17-week gestational age distal femur growth plate demonstrating widespread expression, particularly in the bone (B), proliferating zone of the growth plate (P), and the perichondrium (arrowheads). Non-reactive antibody was used as a control (Ab control). R, reserve zone. H, hypertrophic zone. Scale bars, 100 μm. (B) Picrosirius red staining of the femoral growth plates of a control and the ATD R05-365A fetus. The control growth plate shows normal architecture with clearly distinguishable reserve (R), proliferating (P) and hypertrophic (H) zones. A profound disruption of growth plate architecture is obvious in the GRK2-/- growth plate, which shows irregularly-shaped reserve zone chondrocytes, a lack of discernable proliferating chondrocytes and a significantly under-developed hypertrophic zone. Scale bars, 100 μm. (C) Shortened long bones in the chicken wings injected with CMPD101. The right limb bud was injected with CMPD101, the left one was left as a control. Note the shortened humerus (H), ulna (U) and radius (R) in the CMPD101-injected wing. Mean ± SEM. Mann-Whitney U test; number of animals is indicated. Scale bar, 150 μm. (D) Alcian blue staining of the femoral growth plates of a control and the ATD R05-365A fetus. Note the significantly lesser blue staining of the R05-365A growth plate stained by alcian blue pH 1.0, suggesting impaired sulfation of the extracellular matrix. Scale bars, 200 μm. (E) Alcian blue staining of the chicken limb bud micromasses treated with GRK2 inhibitor CMPD101. Note the gradual inhibition of the cartilage nodules with increasing concentration of CMPD101, quantified below. Mean ± SEM. Mann-Whitney U test; number of micromasses is indicated. A representative experiment of four (pH 2.5) and three (pH 1.0) is shown. Scale bar, 100 μm. (F) Alcian blue staining of the micromasses generated from Grk2+/+ and Grk2-/- NIH3T3 cells. Note the mildly weaker staining by alcian blue pH 2.5 and the nearly absent sulfation stained by alcian blue pH 1.0 in the Grk2-/- micromasses. Central band, median. Box, 1st-3rd quartile. Whiskers, 10%-90% percentile. Mann-Whitney U test; number of micromasses is indicated. Scale bar, 1 mm.
Figure 2A A section of a UMOD filament is shown that consists of 5 consecutive subunits (UMOD 1-5) related by the helical symmetry operation indicated in the top panel. In the middle panel, where the helical axis is represented by a large black arrow, subunits are depicted in cartoon (UMOD 1, 3 and 5) and surface (UMOD 2, 3 and 4) representation to highlight protein-protein interfaces (with UMOD 1 ZP-N and UMOD 5 ZP-C omitted for clarity). In the filament, the ZP-N/ZP-C linker of each molecule (for example, UMOD 3) wraps around the ZP-C domain of the subunit that precedes it (UMOD 2) and the ZP-N domain of the subunit that follows it (UMOD 4); additionally, the ZP-N and ZP-C domains of the same molecule are engaged in interactions with the ZP-C domain of the subunit that in turn precedes UMOD 2 (UMOD 1) and the ZP-N domain of the subunit that follows UMOD 4 (UMOD 5), respectively. As summarized in the bottom panel, every UMOD subunit is thus interacting with another four by being engaged in six interfaces that belong to three different types (ZP-N/ZP-C, black arrow; ZP-N/β1, dark red arrow; E'FG,α1β/ZP-C, light red arrow). Subunits 3, 2 and 4 in this figure correspond to Fig 3 chains A, B and C, respectively. B Path of the interdomain linkers of UMOD1-5, whose domains are outlined in the background. The view is rotated by ~40° around the Y axis, compared to panel A.
Figure 6Healthy control (filled circles) or BLM-induced mice were treated with vehicle (filled squares), 1 nmol (empty circles), 3 nmol (empty squares), or 10 nmol (filled triangles) FA-TLR7-54 and then sacrificed on day 21 for analysis. (A) Analysis of body weight change versus time (n=10). Healthy control (filled circles) or BLM-induced mice were treated with vehicle (filled squares), 1 nmol (empty circles), 3 nmol (empty squares), or 10 nmol (filled triangles) FA-TLR7-54 and then sacrificed on day 21 for analysis. (B) Analysis of the number of cells per milliliter of BALF (n=5). Healthy control (filled circles) or BLM-induced mice were treated with vehicle (filled squares), 1 nmol (empty circles), 3 nmol (empty squares), or 10 nmol (filled triangles) FA-TLR7-54 and then sacrificed on day 21 for analysis. (C) Quantitation of total hydroxyproline content per right lung (Healthy control, n=5; others, n=7-9). Healthy control (filled circles) or BLM-induced mice were treated with vehicle (filled squares), 1 nmol (empty circles), 3 nmol (empty squares), or 10 nmol (filled triangles) FA-TLR7-54 and then sacrificed on day 21 for analysis. (D) H&E staining and trichrome staining of lung tissue (scale bars, 200 μm).
Blocking de novo DNA methylation during adult hippocampal neurogenesis impairs hippocampus-dependent learning and memory.Figure 7A, Mice with adult NSPC-specific deletion of Dnmt3a and Dnmt3b (KO) and controls (WT) were tested in the Morris water maze three months after administration of tamoxifen (n = 13 mice, WT; n = 23 mice, KO). B, Protocol for the Morris water maze test. C, Average distance animals swam per day to find the hidden platform. D, Average time animals needed to reach the platform per day. Note that KO mice showed increased path length and latency on days 4 and 5. E, Time mice spent on day 5 in the quadrant in which the goal was located during the acquisition phase. F, Heatmaps depicting the percentage of individual search strategies used by WT and KO mice per trial (left). Color scheme for the classification into spatial and non-spatial search strategies (right). G, Odds ratio of strategy use in KO compared to WT mice separated by acquisition and reversal phase (p-values were determined by linear regression).
Figure 7A. Workflow for profiling mRNAs bound by the Ncl-Kif5a complex. Nucleolin (Ncl) and Kif5a-binding RNA was isolated from wild type (WT) adult mouse sciatic nerve axoplasm by immunoprecipitation; in addition, DRG neurons from adult WT and GAR+/- mice were cultured in modified Boyden chambers and RNA was isolated from cell body and axonal sides. RNA-seq analysis from the resulting four datasets yielded 11,771 overlapping transcripts. The latter were further processed into a subset of 488 transcripts enriched in Ncl and Kif5a pulldown and depleted in axons of nucleolin GAR+/- mice compared to WT (B). B. Clustering of 488 Ncl/Kif5a-enriched transcripts reduced in GAR+/- versus WT axons. Yellow cluster - transcripts not significantly enriched in the soma of GAR+/- DRG neurons versus the WT control, Purple cluster - transcripts enriched in the soma of GAR+/- DRG neurons versus the WT control. Heatmap shows mean log2 fold changes across four datasets (see also Fig. EV3A), from left to right: 1) nucleolin-binding mRNAs in mouse sciatic nerve axoplasm; 2) Kif5a-binding mRNAs in mouse sciatic nerve axoplasm; 3) mRNA abundance in DRG neuron cell bodies of GAR+/- mice relative to abundance in WT mice; 4) mRNA abundance in DRG neuron axons of GAR+/- mice relative to abundance in wild type mice. All transcripts chosen for this cluster analysis showed significant enrichment in Ncl IP and reduction in GAR+/- axons versus WT, as determined by Rank-Rank Hypergeometric Overlap (RRHO) as well as a fold-change in Kif5a IP versus control > 2. C. Genes comprising the purple cluster - transcripts significantly enriched by both Ncl and Kif5a immunoprecipitation and showing a reduction in GAR+/- axons concurrent with an enrichment in GAR+/- soma (compared to the WT control). Inpp5f (highlighted in red) was chosen for follow-up. D. Representative images for FISH analysis of Inpp5f in DRG neurons treated for 48h with 10 µM AS1411 or 10 µM control aptamer, replated and grown for 18h. FISH signal is shown in gray, cell somata (left) and axons (right) are visualized by neurofilament immunostaining (magenta). Scr - scrambled FISH probe served as a negative control. Scale bar - 10 µm. E. Quantification of axonal Inpp5f signal in (D) as density of RNA granules along axons. n = 11-19 cells per sample; means ± SEM; * p<0.05, unpaired Student's t-test. F. RT-qPCR analysis of axonal Inpp5f mRNA levels in DRG neurons grown in Boyden chambers and treated for 48h with 10 µM AS1411 or 10 µM control aptamer. Axon/cell body ratios of Inpp5f normalized to Gapdh levels are shown. n = 4 independent biological repeats; means ± SEM; * p<0.05, unpaired Student's t-test.
Dyn2 knockdown disrupts autophagic lysosomal reformation (ALR) and lysosomal tubule scission.Figure 5.Hep3B cells treated with either a nontargeting control siRNA (A and B, siNT) or an siRNA targeting human Dyn2 (C and D, siDyn2) were fixed and stained with an antibody specific for LAMP1 after a 24-h starvation period in media containing 0.1% FBS. Control cells displayed normal lysosomal compartments, with LAMP1-stained structures dispersed throughout the cytosol, some containing short reformation tubules (B′). After Dyn2 knockdown, many cells contained enlarged autolysosomes with a noticeable increase in persistent and lengthy reformation tubules emanating from LAMP1-positive compartments (C′ and D′). Bars: (A-D) 20 µM; (A′ and B′) 5 µM; (C′ and D′) 10 µM. (E and F) Measurements of the length (E) and distribution (F) of LAMP1-stained ALR tubules in control (siNT) and siDyn2-treated cells. Data represent the mean derived from measurements of tubules in a minimum of 40 cells for each experimental group over 6 independent experiments. Error bars represent SE; *, P ≤ 0.05; **, P ≤ 0.01. (G) TEM of siDyn2-treated Hep3B hepatocyte exhibiting enlarged autolysosomal structures with extensive tubulation (arrows). Bar, 2 µm. (G′) Inset showing enlargement of autolysosomal tubule (arrows). Bar, 1 µm. (H) Tubulating autolysosome with engulfed lipid droplet. Bar, 0.5 µm.