EMBO/sd-character-level-ner huggingface dataset:
Figure 1Schematic diagram of suction blistering and sample collection. A blister produced on C57BL/6 wild-type (WT) mouse dorsal skin at P1. BM: basement membrane. Blistered samples at P1. (B) Hematoxylin and eosin (H&E, top) and alkaline phosphatase (AP, bottom) staining. Hair follicles (HFs) remaining in the dermis (indicated by arrows). Scale bar: 500 μm. (C) α6 Integrin (ITGA6, indicated by arrowheads) and type IV collagen (COL4, arrows) labeling (left). Laminin 332 (L332, arrows) staining (right). Scale bar: 100 μm. (D) Ultrastructural findings of blistered skin (left image: blister roof, right image: blister bottom) . Hemidesmosomes (white arrowheads) and lamina densa (arrows) are indicated. Scale bar: 1 μm. H&E (left), pan-cytokeratin (PCK, middle), and ITGA6 staining (right) at P2. The regenerated epidermis is indicated by arrowheads. Scale bar: 200 μm. Keratin 14 (K14) and keratin 10 (K10) staining of the nonlesional (intact) and lesional (blistered) skin at P2 (upper images and inlets: sections, lower images: whole-mount imaging). HFs are indicated by arrows in the whole-mount images. Scale bar: 30 μm. H&E (top) and loricrin (LOR, bottom) staining at P3. Scale bar: 200 μm. H&E (left), PCK (middle), and LOR staining (right) at P4. Scale bar: 200 μm.
Combinatorial treatments induces CLEAR network associated with a blockage in autophagy A Relative mRNA expression levels of indicated CLEAR network genes in HCT116 cells treated as indicated ((NEN (N) 1.Figure 3A Relative mRNA expression levels of indicated CLEAR network genes in HCT116 cells treated as indicated ((NEN (N) 1.2 µM; Domperidone (Domp, D), Imipramine (Imi), Desipramine (Desi) and Amitriptyline (Ami), each 30 µM; Clomipramine (Clomi) 20 µM), determined by qPCR. Data information: In (A) (first (TFE3) and third (CD68) graph (N=3); second (TFE3) and fourth (CD68) graph (N=5, besides NEN+Clomi (N=4) and NEN+Desi (N=4)) data are presented as mean (SD) and were analyzed by a one-way ANOVA with Tukey post-hoc test. In the second and the fourth graph, significance is indicated for the comparison of combinatorial treatments (NEN+TCAs) to controls (Ctrl). * p< 0.05; ** p< 0.01;*** p< 0.001; **** p < 0.0001. B Immunofluorescence (IF) of HCT116 cells treated with DMSO (Control (Ctrl)), NEN, Domperidone (Domp) or NEN+Domp as indicated (NEN, 1.2 µM; Domp, 30 µM) for 16 h and stained with anti-Lamp1 (top panel), Alexa-555 LC3 (middle panel) co-stained with Alexa-647 p62 (bottom panel). Nuclei, Blue (dapi). Scale bar 20 µm. Mean (SEM) fluorescence of Lamp1 (Ctrl, NEN, Domp (N=20 cells); NEN+Domp (N=24 cells)), LC3-II (Ctrl, NEN+Domp (N=16 cells)); NEN, Domp (N=16 cells)) or P62 (Ctrl (N=38 cells); NEN (N=27 cells); Domp (N=28 cells); NEN+Domp (N=21 cells)) was quantified and analyzed by one-way ANOVA with Tukey post-hoc test (for LC3-II) or Kruskal-Wallis with Dunn´s post-hoc test (for Lamp1 and p62). * p< 0.05; ** p< 0.01;*** p< 0.001; **** p < 0.0001. C Immunoblot analysis and quantification (mean ± SD) of LC3-II/LC3-I ratios and P62 in HCT116 cells. Cultured cells were treated with DMSO (Control), NEN or NEN and Domperidone (Domp) as indicated (NEN, 1.2 µM; Domp, 30 µM) for 16 h. Protein band signal intensities were normalized to vinculin (loading control) and the ratios LC3-II bands LC3-I (N=3) and p62 levels (N=3) were analyzed by one-way ANOVA with Tukey post-hoc test. * p< 0.05; *** p< 0.001; **** p < 0.0001. D Immunoblot analysis and quantification of LC3 in HCT116 cells (mean ± SEM). Cultured cells were treated with DMSO (Control (Ctrl)), NEN or NEN and Domperidone (NEN+Domp) as indicated (NEN, 1.2 µM; Domp, 30 µM) for 16 h, followed by control or 20 µM chloroquine treatment for 3 h. Protein band signal intensities were normalized to vinculin (loading control) and the ratios LC3-II bands in presence or absence of chloroquine treatment (N=6) were analyzed by Kruskal-Wallis with Dunn´s post-hoc test. * p< 0.05; *** p<0.001.
Figure 2(A) Schematic diagram for the induction of CiPSCs from MEFs using the "three-step" method (Zhao et al., 2018). (B) Durations of dox addition at different time points during chemical induction (upper) and assessment of AP+ colonies (bottom). Error bars, mean ± S.D.; n = 3 biological replicates per condition; **P < 0.01, ***P < 0.001 as compared with condition-1, by two-tailed Student's  t-test. n.s., not significant. (C) Morphological changes at distinct time points during D-CiPS induction. Scale bar, 100 μm. (D) Representative images of the number of AP+ positive colonies. n = 5 biological replicates per condition. (E) RT-qPCR analysis of select ZGA-related genes in dox-untreated D-CiPSCs and canonical CiPSCs at passage 1. The value in CiPSCs was set as 1, and data shown are mean expression values relative to GAPDH. Error bars, mean ± S.E.M.; n = 3 biological replicates per group; ***P < 0.001 by two-tailed Student's t-test. (F) Representative immunostaining images of D-CiPSCs expressing pluripotency markers. n = 5 biological replicates. Scale bar, 50 μm. (G) Histology by H&E staining of teratoma tissues derived by D-CiPS cells. n = 3 biological replicates. Scale bar, 200 μm. (H) Chimeric mouse generated from D-CiPSCs. (I) Karyotype analysis of D-CiPSCs and canonical CiPSCs. The majority of D-CiPSCs and CiPSCs cells possessed a normal karyotype. Scale bar, 10 μm. (J) Bisulfite sequencing analysis of demethylation of Oct4 and Nanog promoters in D-CiPSCs or canonical CiPSCs. Filled and empty squares represent methylated and unmethylated CpGs, respectively.
Figure 3A Cumulative number of times that genes were present amongst the top 15 ranking genes from 25 reiterative RF models. B Variable Importance (Gini) for top 20 genes from RF model that was used to define the 15-gene signature. C Box plots showing relative expression levels of top classifier genes in our RT-PCR dataset. Displayed are inverse deltaCt values (zero minus ΔCt), such that higher values indicate higher expression levels for each sample in this study.
Figure 3(A) Total levels of PAR in whole cell lysate were measured using western blot analysis. Wild-type (WT) and Ews-/- mBA cells were treated with H2O2, (1 mM, 20 minutes) with or without Olaparib (5 μM, 7 hours). (B) The kinetics of PAR accumulation was analyzed by western blotting. WT and Ews-/- cells were treated with H2O2 (1 mM, 20 minutes), followed by incubation in fresh media for the indicated times to allow release of DNA damage (C) Immunohistochemistry detection of PAR by ADP-ribose antibody in WT and Ews-/- cells after H2O2 treatment (1 mM, 20 minutes) and recovery from DNA damage. Right graph displays mean of intensities measured from 500 cells. Data represented as mean ±SEMs, significance determined by One-way ANOVA, ***P < 0.001. Scale bar indicate 20 μm. (D, E) Whole cell level of PAR was measured in WT and Ews-/- mBA cells upon treatment with (D) MMS (0.02%, 60 minutes) (E) with or without recovery from DNA damage. (F) NAD+/NADH ratio was measured in mBA cell lines following treatment of H2O2 (1 mM, 10 minutes). Data represented as mean ±SEMs, and technical repeats (n=3). Significance determined by Student's t test, two-tailed, **P < 0.01, ***P< 0.001.
Figure 4(A) WT, and Irgm2-/- BMMs were stimulated overnight with IFNγ or left untreated and cell lysates were collected. Lysates were assessed for Casp11, Gbp2, and actin protein levels via immunoblotting. (B) IFNγ-primed and unprimed WT, Irgm2-/-, Gbpchr3-/- and Aim2-/- BMMs were infected with Francisella novicida (MOI 10) and LDH release measured at 4 hpi. (C) WT, Irgm2-/-, Gbpchr3-/- and Irgm2-/- Gbpchr3-/- BMMs were treated with LPS (1 µg/ml). IL-1β, IL-18, and LDH release were assessed at 24 hpt. (D) IFNγ-primed WT, Irgm2-/-, Gbpchr3-/- and Irgm2-/- Gbpchr3-/- BMMs were treated with LPS (1 µg/ml) for 24 hours and cell lysates and supernatants collected. Protein levels in cell lysates (Caspase-1, and actin) and supernatants (Caspase-1 p20) were visualized via immunoblotting. (E) Model depicting regulation of caspase-11 activation by Irgm2 and Gbps.
IGSF10 protein structure and position of N-terminal mutations.Figure 3A: IGSF10 domains and N-terminal mutations identified in the study. Region I contains leucine-rich repeats (LRR) 1-7 flanked by a LRR N-terminal (LRR Nt) and C-terminal (LRR Ct) cap. Region II is structurally disordered. Region III contains two Ig-like domains (Ig). Region IV is structurally disordered. Region V contains 10 Ig-like domains (Ig). B: Protein tertiary structure as predicted by in silico analysis. C: Biological consequences of the 2 identified N-terminal mutations. Both WT and mutant N-terminal protein fragments (p.Arg156Leu and p.Glu161Lys) were expressed in HEK293 cells as demonstrated by western blotting. The GFP-tagged protein products of both were not detected in the conditioned media of mammalian cells, as compared to wild type (WT), and appear to be retained in the intracellular compartment (mean ± s.e.m.; n = 3). Ponceau redstaining is shown to demonstrate equal protein loading for conditioned media. UT - untransfected negative control; nd - not detected; *: p=0.01094 (WT vs p.Arg156Leu) and p=0.04408 (WT vs p.Glu161Lys) by 2-tailed t-test. It has to date not been possible to test cytoplasmic retention for the two C-terminal mutations due to difficulty expressing the full length or C-terminal protein fragment in mammalian cells.
Inhibition of autophagy potentiates sodium butyrate-induced apoptotic cell death.Fig 10  HCT-116 (A) and HT-29 (B) cells were exposed to 5mM 3-MA or 5μM chloroquine (CQ) for 30 min and then treated with 2mM sodium butyrate (NaB) for 24 h. The percentage of apoptotic cells (annexinV+/PI-) was quantified by flow cytometry using an annexinV/PI assay and expressed as mean ± SD of three independent experiments. One-way ANOVA was used for statistical analysis to compare control cells and NaB treatments. *p<0.05, ** p<0.01 compared to control. (C.D.) Cells were treated as described above and the proteins were harvested for Western blot analysis. Expression of full-length and cleaved PARP in HCT-116 (C) or HT-29 (D) cells was compared to GAPDH, the loading control. The ratio of cleaved PARP to total PARP was calculated and the fold change in normalized PARP expression relative to cells treated with 2mM NaB is shown as the mean ± SD of three independent experiments. One-way ANOVA was used for statistical analysis. *P<0.05, ** p<0.01, compared to the 2mM NaB group.
The gene encoding TP53INP1 is expressed in pancreatic endocrine cells.Figure 3A, B (A, B) Immunocytofluorescent staining of TP53INP1 (red) and insulin (green) in mouse pancreatic sections (A) and single human islet beta cell (B). Scale bars represent 50 μm (A) and 10 μm (B). C-E Quantitative PCR for Tp53inp1 mRNA levels in tissues and cells from rat (C) and C57BL/6Jmice fed with a normal diet (5% fat; ND) or an high-fat diet (45% fat; HFD) (D, E). Results are expressed as the mean ± SEM and are representative of two independent experiments. n = 2 for ratliver, islets and mousespleen; n = 4 for INS-1E cells; n = 5 for mouseexocrine pancreas; n = 6 for Min6 cells and ND and HFD islets; n = 11 for mouseislets. *P = 0.035 for HFD versus ND.
Defective recruitment of ERCC1 to sites of DNA damage in USP45 knockout cellsWild‐type (WT), USP45 knockout (KO) or USP45 KO cells re‐expressing wild‐type USP45 (RESWT) or catalytically inactive USP45 [Cys199Ala] (RESCA) were used to analyse ERCC1 foci formation.Figure 6A Wild‐type (WT), USP45 knockout (KO) or USP45 KO cells re‐expressing wild‐type USP45 (RESWT) or catalytically inactive USP45 [Cys199Ala] (RESCA) were used to analyse ERCC1 foci formation. Staining and analysis of endogenous ERCC1 foci formation was undertaken before (left panel), or after DNA damage induction by mitomycin C (40 ng/ml, 16 h, middle panel) or UV (20 J/m2 followed by 3‐h recovery, right panel). Three independent experiments were performed in which 500 cells per experiment were analysed. Scale bar, 10 μm.B Proportion of cells displaying more than 10 endogenous ERCC1 foci were quantified. Three independent experiments were performed in which 500 cells per experiment were analysed. Results are the mean of 3 experiments ± SD.C The wild‐type and USP45 knockout U2OS cells were treated with mitomycin C (40 ng/ml, 16 h) and total number of ERCC1 staining foci per cell was quantified in at least 50 independent cells at the indicated times. Similar results were obtained in two separate experiments. The data are presented as the average number of ERCC1 staining foci per cell. D As in (C), except that γ‐H2AX staining foci were analysed. The data are presented as the average number of γ‐H2AX staining foci per cell.
Immunogenicity and protective efficacy of PiCoVacc in nonhuman primates.Fig. 3primates.Macaques were immunized three times through the intramuscular route with various doses of PiCoVacc or adjuvant only (sham) or placebo (n=4). SARS-CoV-2-specific IgG response (A) and neutralizing antibody titer (B) were measured. Points represent individual macaques; dotted lines indicate the limit of detection; horizontal lines indicate the geometric mean titer (GMT) of EC50 for each group. Protective efficacy of PiCoVacc against SARS-CoV-2 challenge at week 3 after immunization was evaluated in macaques (C-F). Viral loads of throat (C) and anal (D) swab specimens collected from the inoculated macaques at day 3, 5 and 7 pi were monitored. Viral loads in various lobe of lung tissue from all the inoculated macaques at day 7 post infection were measured (E). RNA was extracted and viral load was determined by qRT-PCR. All data are presented as mean ± SEM. Histopathological examinations (F) in lungs from all the inoculated macaques at day 7 post infection. Lung tissue was collected and stained with hematoxylin and eosin.
Kdm3b plays an important role in reprogramming to pluripotency a.Figure 1Quantification of NANOG-positive iPSC colonies obtained after 14 days of OSKM induction in MEFs treated with either non-target or Kdm3b siRNA in FBS media. Values represent fold change over non-target control providing relative quantitation. Quantification of NANOG-positive iPSC colonies obtained after 12 or 14 days of OSKM induction in MEFs treated with either non-targeting or Kdm3b siRNA in KSR media. Values represent fold change over non-target control providing relative quantitation. Quantification of NANOG positive colonies after retroviral expression of OSKM (Left) or OSK (Right) in MEF treated with non-targeting or siRNA targeting Kdm3. Nanog counts were performed on Day 16 for OSKM or Day 21 for OSK reprogramming. Values represent fold change over non-target control providing relative quantitation. Immunoblot of H3K9me1/2 histone demethylase in WT or Kdm3b CRISPR targeted pre-iPSC clones. These pre-iPSCs carry a Nanog-GFP reporter. Parent line (WT), and two Kdm3b-KO clones using gRNA targeting either exon 1 (KO#1) or exon 2 (KO#2) are shown. RNA Polymerase II levels are the loading control. Quantification of Nanog-GFP positive cells after 10 days of AA+2i treatment of WT or Kdm3b-deleted pre-iPSCs. Immunoblot of H3K9 methylation levels upon addition of H3K9 methyltransferase inhibitor UNC0638 in WT or Kdm3b-KO pre-iPSCs. Quantification of percent Nanog-GFP positive cells after 10 days of AA+2i or AA+2i with 5µM UNC0638 in WT or Kdm3b deleted pre-iPSCs.
Immediate late genes (ILGs) translate signal duration into response amplitude.Figure 6 Figure 6 - Immediate late genes (ILGs) translate signal duration into response amplitude. A: Upper panel: pERK2 log2 fold changes upon different input scenarios (sustained: 4OHT; two-hour pulse: 4OHT+U0126). Lower panel: Response amplitude across temporal clusters and signal durations. Bold lines show median cluster amplitude at each time point. B: Capacity to decode signal duration. ILGs translate ERK signal duration into response amplitude. IEGs are only partially able to do so, as many of these genes are still strongly induced upon shortened signal durations. C: qPCR validation to test different ERK signal durations. HEK293∆RAF1:ER cells were treated with 4OHT and U0126 for different periods of time to generate signal duration scenarios of 0.5 to 8 hours (cf. Fig EV1 C). mRNAs of IEGs EGR1 and FOS relay signal duration to response duration, whereas ILGs CLU and FOSL1 decode signal duration to response amplitude (qPCR data for all 17 validated mRNAs is shown in Fig EV6 A). D: Relation between signal duration and response amplitude for IEGs and ILGs derived from qPCR validation data. Median amplitude is based on six qPCR-validated ILGs and six qPCR-validated IEGs. E: Quantification of western blots to present protein log2 fold changes of sample genes upon sustained ERK signalling (4OHT-induced) and transient ERK signalling (EGF-induced) in HEK293∆RAF1:ER cells. F: Conservation of signal duration decoding to response amplitude in two prominent model systems for ERK signal duration: NGF and HRG cause more sustained ERK signalling compared to EGF treatment in PC12 and MCF7 cells respectively. Decoding of signal duration to response amplitude is clearly governed by ILGs.
Figure 1A, B. RT-qPCR analysis of Zfp207 and Oct4 in mouse ESCs during (A) retinoic acid (RA)-induced and (B) embryoid body (EB)-mediated differentiation. Nestin was used as a neuronal differentiation marker to monitor RA-mediated differentiation. mRNA levels are relative to the expression at day 0. C, D. Western blot of ZFP207 and OCT4 of mouse ESCs during (C) RA-induced and (D) EB-mediated differentiation. E, F. (E) RT-qPCR and (F) western blot to monitor the knockdown efficiency of Zfp207 (sh1 and sh2) (shScr). G. Cell proliferation rate of shScr, sh1 and sh2 ESCs assessed over a period of 6 days. H. Percentage of live (Annexin V-) and apoptotic cells (Annexin V+) in sh1 and sh2 mouse ESCs compared to shScr. I. Bar chart displaying the cell cycle distribution in sh1 and sh2 mouse ESCs relative to shScr. J, K. (J) AP staining of shScr and Zfp207-depleted (sh1 and sh2) mouse ESCs. Scale bars, 20 µM. (K) Percentage of fully differentiated (FD), partially differentiated (PD) and undifferentiated (UN) ESC colonies in shScr, sh1 and sh2. L. Immunofluorescence analysis of SSEA1 in shScr, sh1 and sh2 ESCs. DAPI was used as the nuclear marker. Scale bars, 20 μm. M, N. (M) Western blot of ZFP207, OCT4, NANOG, and SOX2 in shScr, sh1 and sh2 ESCs and (N) RT-qPCR analysis of Oct4, Nanog, and Sox2 in shScr, sh1 and sh2 ESCs; data is relative to shScr. (O) Western blot of OCT4 during a 9-h cycloheximide (CHX) time course treatment in shScr, sh1 and sh2 ESCs. (P) Protein degradation curves were made after quantification and normalization of the bands from (O). (Q) Western blot of OCT4 during 4 and 8 h treatment with the proteasome inhibitor MG132. (R) graph bars after quantification and normalization of the bands from (Q).
| RNR inhibition overcomes the SAMHD1-mediated barrier to ara-C in AML mouse models Kaplan-Meier analysis of NOD/SCID mice injected i.Figure 2Kaplan-Meier analysis of NOD/SCID mice injected i.v. with luciferase-expressing SAMHD1+/+ or -/- THP-1 cell clones (day 0) and treated with ara-C and/or HU as indicated (day 6). n = 6 per treatment group. Kaplan-Meier analysis of NOD/SCID mice injected i.v. with luciferase-expressing SAMHD1+/+ or -/- HL-60/iva cell clones (day 0) and treated with ara-C and/or HU as indicated (day 6). n = 6 per treatment group. Kaplan-Meier analysis of NOD/SCID mice injected i.v. with luciferase-expressing SAMHD1+/+ THP-1 cell clone (day 0) and treated with ara-C and/or dF-dC as indicated (day 6). n = 7 per treatment group. Kaplan-Meier analysis of CD45.2 C57BL/6J mice injected i.v. with murine MLL-AF9-transformed AML blasts (day 0) and treated with ara-C and/or HU days 20-24. n = 5 per treatment group, except for vehicle (n = 4).
Rolling mediates inter-layer coupling and facilitates pore opening.Figure 4iMODfit fitting of the GluN1/GluN2B inhibited state into the cryo-EM map EMD-3352 (target structure in a putatively active state; TMD not resolved). Left, Evolution of the RMSD calculated against PDBs 5FXG (putative active state), 5FXI (non-active state) and 5IOV (inhibited state) over 2267 aligned Cα. The trajectory is divided in three steps (vertical dashed lines). Right, illustration of the fitting by superposition of the target EM density map (envelope representation) and the initial and final frames of the trajectory (line representation). GluN1 and GluN2B subunits are colored red and blue, respectively Left, evolution of selected collective variables during the iMODfit simulation. Each plot illustrates two collective variables, each with its own Y-axis (black or red). The upper panel (B) focuses on the NTD region (distance between the two GluN1 NTDs at the "apex" of the receptor; volume of the ifenprodil binding pocket at the interface between GluN1 and GluN2B NTDs), the middle panel (C) on the distance between NTD lower lobes and the ABD rolling motion, and the lower panel (D) on the ABD-TMD connection and the M3 channel gate dilation. Right, illustration of the three steps of the fitting showing in step 1 (frame 1 to 8) the reduction of the inter NTD dimer distance; in step 2 (frame 8 to 15) the NTD lower lobes getting closer and the associated rolling of the ABD dimers; in step 3 (frame 15 to 22) the distance changes in the ABD-M3 linker and the channel gate radius. The inset highlights a top view of the pore lining helixes M2 (P-loop) and M3 with a same-size circle emphasizing pore dilation during step 3. The GluN1 linker distance represents the distance between the center of masses (COM) of both GluN1-R663 residues and the COM of the M3 SYTANLAAF sequence residues, GluN2B linker distance represents the distance between both GluN2B-E65