EMBO/sd-character-level-ner huggingface dataset:
Figure 5Yeast three-hybrid assays showing that CBF1 represses the interaction of the Pfr-phyB with PIF4 in yeast cells. AD-PIF4, phyB-BD and CBF1 were expressed in the yeast strain Y190 as indicated. The β-galactosidase activities were measured by liquid culture assays using ONPG as the substrate. Error bars represent SD of three independent yeast clones. Yeast three-hybrid assays showing that CBF1 represses the interaction of the Pfr-phyB with PIF5 (B) in yeast cells. AD-PIF4, phyB-BD and CBF1 were expressed in the yeast strain Y190 as indicated. The β-galactosidase activities were measured by liquid culture assays using ONPG as the substrate. Error bars represent SD of three independent yeast clones. LCI assays showing that CBF1 inhibits phyB interaction with PIF4 (C) in plant cells. Different letters represent significant differences by one-way ANOVA with Duncan's post hoc test (P < 0.05). LCI assays showing that CBF1 inhibits phyB interaction with PIF5 (D) in plant cells. Different letters represent significant differences by one-way ANOVA with Duncan's post hoc test (P < 0.05). Semi-in vivo pull-down assays showing that CBF1 inhibits phyB interaction with PIF4 (E) The total proteins extracted from 4-d-old 35S:phyB-GFP seedlings grown at 22°C in continuous R light served as the bait, and GST-PIF4 fusion proteins served as the prey. Equivalent amounts of phyB-GFP protein extract and GST-PIF4 fusion proteins, and increasing amounts of His-CBF1 were added as indicated before anti-GFP IP assays were performed. The pulled-down proteins were subjected to immunoblotting with antibodies against phyB, GST and His, respectively. Numbers below the anti-GST blots indicate the relative band intensities of co-precipitated GST-PIF4 normalized to those of precipitated phyB-GFP, respectively. The ratio of the first clear band was set to 100 for each blot. Semi-in vivo pull-down assays showing that CBF1 inhibits phyB interaction with PIF5 (F). The total proteins extracted from 4-d-old 35S:phyB-GFP seedlings grown at 22°C in continuous R light served as the bait, and GST-PIF5 fusion proteins served as the prey. Equivalent amounts of phyB-GFP protein extract and GST-PIF5 fusion proteins, and increasing amounts of His-CBF1 were added as indicated before anti-GFP IP assays were performed. The pulled-down proteins were subjected to immunoblotting with antibodies against phyB, GST and His, respectively. Numbers below the anti-GST blots indicate the relative band intensities of co-precipitated GST-PIF5 normalized to those of precipitated phyB-GFP, respectively. The ratio of the first clear band was set to 100 for each blot.
Figure 6(A) Summary and representative immunoblots of PKCα expressions in mouse bone marrow neutrophils from PKCα-/-, PKCα+/-, and their littermate WT (PKCα+/+) mice. Vinculin served as loading control. (B) Representative immunoblot (IB) detection of the phospho-lamin B (Ser) and total lamin B with lamin B protein purified by immunoprecipitation (IP) with anti-lamin B from mouse peritoneal mPMNs from WT vs PKCα-/- mice, stimulated without or with PAF for 3 h. (C) Summary analyses of NET formation in peritoneal mPMNs, from WT, PKCα+/-, or PKCα-/- mice, which were treated without or with either 5 µM or 10 µM PAF for 3 h following by 2% PFA fixation and fluorometric microplate analysis. (D) The endpoint analysis of NET-DNA release index was detected by coincubation of primary mouse peritoneal mPMNs from WT and PKCα-/- mice without (control) or with 10 µM PAF in medium containing 1 μM Sytox Green dye with recording by a microplate reader at the end of the 3 h time point. The NET-DNA release index was reported in comparison to an assigned value of 100% for the total DNA released by neutrophils lysed by 0.5% (v/v) Triton-X-100. (E, F) (E) Representative and (F) summary analysis of NET formation of mouse peritoneal mPMNs from WT vs PKCα-/- mice stimulated without or with 10 µM PAF for 3 h, and stained with both cell permeable SYTO Red dye and cell impermeable Sytox Green dye, without fixation. Images were taken by Olympus confocal microscopy, followed by automated quantification of NETs on 5-6 non-overlapping area per well using ImageJ for calculation of % cells with NET formation. (E) Scale bars, 50 μm. % cells with NET formation by image analysis using ImageJ (F)
Figure 7Schematic illustration of immunization schedule with complex InsA peptides (cInsA) intraperitoneally and insulin-specific protective IgM (PR-IgM) in 48 hours cycles intravenously (i.v.). *monitoring: diabetes symptoms were only observed within cInsA only group. Blood and urine glucose levels of mice immunized with complex InsA-peptides (cInsA) (red, n=5) and cInsA plus intravenously injected (i.v.) pIgM (salmon, n=5) at day 7. Dots represent individual mice. Mean ± SD, statistical significance was calculated by using Mann-Whitney-U test, *P < 0.05. Blood glucose levels of mice intravenously injected with total IgG pulldown of cInsA immunized mice (red, n=4), or total IgG (cInsA) together with anti-insulin PR-IgM (black, n=4). Measurements were done 5 hours post intravenous injection. Data were collected from two independent experiments. Mean ± SD, statistical significance was calculated by using Mann-Whitney-U test, **P < 0.01. (D) Serum (dilution: 1:50) and (F) insulin-specific IgM (concentration: 500 ng/mL) tested for dsDNA (left panel, coating: calf thymus dsDNA) and insulin (right panel, coating: Insulin) reactivity measured by ELISA. Mice were immunized with cInsA and serum was collected on day 7 (D: n=8, left; n=5, right; F: n=4) and day 85 (D: n=4, left; n=5, right, F: n=4) post immunization. Dots represent individual mice. Mean ± SD, statistical significance was calculated by using Mann-Whitney-U test, **P < 0.01, ****P < 0.0001. Anti-nuclear-IgM (ANA) of control-immunized (CI, n=3) and complex InsA peptide (cInsA) immunized mice on day 7 (n=3) and day 85 (n=3) analyzed by indirect immunofluorescence. We used (E) total serum (dilutions: 1:20) or (G) Insulin-specific IgM (concentration: 500 ng/mL) purified from serum at indicated days on commercial HEp-2 slides. Scale bar: 10 µm. Green fluorescence indicates IgM bound to nuclear structures. Data are representative for three independent experiments with isotype control: n=3, day 7: n=3, day 85: n=3. Coomassie stained SDS-page showing primary (cInsA d7) and memory (cInsA d85) Insulin-specific purified IgM pre-incubated with Insulin and calf thymus dsDNA. Samples were loaded onto the gel after size exclusion with a cut-off at 10.000 kD (fractions referring to >/< 104 kD). IgM heavy chain (HC): 69 kD, IgM light chain (LC): 25 kD, J-chain: 15 kD. Data shown are representative of three independent experiments. Blood glucose levels of mice intravenously injected with either IgM isotype ctrl (n=6), anti-insulin PR-IgM (d85, n=5), anti-insulin primary IgM (d7, n=4) after Insulin-specific pulldown. Mean ± SD, statistical significance was calculated by using repeated measure ANOVA test showing comparison of red and black line, ***P < 0.001, ****P 0.0001.
Copper-binding conformations and PrP octarepeat sequence mutations.Figure 1A The structure of full-length PrP and derivative C1 and C2 fragments with an enlarged view of the cleavage sites is shown. The C2 (β) site was mapped in HpL3-4 cells by (Mange et al, 2004). Ragged C1 (α) cleavage sites in brain are shown (Chen et al, 1995), and a dotted arrow indicates that additional sites have been mapped rightward of this position in vitro (McDonald et al, 2014). Ragged N-terminal cleavage sites of PrP27-30 from prion-infected mousebrain are indicated by small arrows (Howells et al, 2008). The positions of antibody epitopes are also shown (residues in brackets). Grey shading reflects the minimum length for a C2 fragment defined operationally by reactivity with 12B2 antibody. B S1 and S3 alleles, by virtue of limited geometries (modified from Chattopadhyay et al, 2005), may have access to distinct subsets of the full range of biological activities of WT PrP, which has access to interchangeable component 1, 2 and 3 geometries. C Sequences of synthetic PrP alleles differing in the octarepeat region of mousePrP. Conserved histidines in octarepeats 2-5 are coloured blue and missense mutations are shown in red. Overproduction of C2 fragment is indicated by ++ signs to the right of the panel.
DCV release probability upon different stimulations A Schematic representation of NPY-pHluorin as optical reporter for DCV fusion.Figure 5Schematic representation of NPY-pHluorin as optical reporter for DCV fusion. Inset below shows F/F0 of single DCV fusion event. Scale bar: 1 µm. Zoomed confocal image of NPY-pHluorin expressing (green) hippocampal neuron (DIV14) immunostained for ChgB (red) and corresponding fluorescence intensity plot showing overlap between NPY-pHluorin and ChgB. Scale bar: 2 µm. Manders' coefficients of colocalization between NPY-pHluorin and ChgB per neuron. Total ChgB puncta per neuron in uninfected (control) and NPY-pHluorin infected neurons (NPY-pH). Student's t-test: n.s. = not significant, p = 0.90. Mean ChgB fluorescence intensity per puncta per neuron in uninfected (control) and NPY-pHluorin infected neurons (NPY-pH). Student's t-test: n.s. = not significant, p = 0.71. Correlation between ChgB puncta per neuron and total neurite length (mm) for uninfected control (grey) and NPY-pHluorin infected (green) neurons. Linear regression, inset shows goodness of fit (r2) and significance (p-value) per group. Slopes are not significantly different, ANCOVA: p = 0.19. Single isolated hippocampal neurons (DIV14) were labeled with NPY-pHluorin. DCV fusion events were quantified per neuron upon different stimulation paradigms. Histogram of DCV fusion events per stimulation per time point. Stimulations include 1 action potential (AP) (grey), 16 bursts of 50 AP at 50 Hz (light-green), superfusion with 60 mM KCl Tyrode solution for 30 s. (dark-green) and superfusion with 5 μM Ionomycin for 30 s. (blue). Inset: Typical ∆F/F0 trace of rise in intracellular calcium (Fluo5-AM) upon different stimulations. Traces are corrected for baseline (first 10 frames) and normalized. Single isolated hippocampal neurons (DIV14) were labeled with NPY-pHluorin. DCV fusion events were quantified per neuron upon different stimulation paradigms. Cumulative plot of fusion events per neuron for different stimulations. Single isolated hippocampal neurons (DIV14) were labeled with NPY-pHluorin. DCV fusion events were quantified per neuron upon different stimulation paradigms. DCV fusion events per neuron per stimulation. Kruskal-Wallis with Dunn's correction: *p < 0.05, **p < 0.01, ***p < 0.001. 1AP vs. 5 μM Ionomycin, 16x50 AP,50Hz vs. 60 mM KCl and 60 mM KCl vs 5 μM Ionomycin were non-significant, p > 0.05. DCV release probability per neuron per stimulation. Kruskal-Wallis with Dunn's correction: **p < 0.01, ***p < 0.001. 1AP vs. 5 μM Ionomycin, 16x50 AP,50Hz vs. 60 mM KCl and 60 mM KCl vs 5 μM Ionomycin were non-significant, p>0.05. DCV release probability per neuron in CaMKII-positive and negative hippocampus neurons (DIV14) upon repetitive electrical stimulation (16x50AP at 50 Hz). Mann-Whitney U test: n.s. = not significant, p = 0.12.
Figure 2(A) Differential gene expression analysis of H37Rv treated with Q203, ND-011992, or combination (H37Rv-Combo), and ∆cydAB treated with Q203 compared to untreated controls. The log2-fold differences in gene expression of various conditions relative to the untreated control is indicated using a sliding scale where higher expression is reflected in red and lower expression is reflected in blue, with a midpoint signifying no difference in white. (B) Effect of cydABDC over-expression on the Oxygen Consumption Rate (OCR) and intracellular ATP levels. in M. bovis BCG treated with ND-011992 in the presence of 100 nM Q203. IC50: Inhibitory Concentration 50%. (C) Absence of the Cyt-bcc:aa3 sensitized H37Rv to ND-011992. M. tuberculosis H37Rv wildtype (Blue circles), ∆ctaE-qcrCAB (red squares) and complemented strain (green triangles) were treated with ND-011992. Growth inhibitory potency (MIC50) was recorded after 2 to 3 weeks of incubation. (D) The reduced minus oxidized difference spectrum of M. smegmatis ΔqcrCAB inverted membrane vesicles (IMVs). The green line represents the spectrum without treatment, while the blue line represents the spectrum of the IMVs in the presence of 10 μM ND-011992.
Both sporozoite surface and EEF vacuolar membrane antigens are susceptible to vaccine-induced CD8+ T cells for killing, resulting in sterile protection.Figure 6(A) Schematic overview of experimental design. Mice received 2x106 OT-I splenocytes, and one day later they were either vaccinated with 1x108 ifu recombinant AdHu5 expressing whole ovalbumin (AdOVA) (n=5 per challenge group) or left untreated (n=4 per challenge group). (B, C) Flow cytometry (B) and scatter plots (C) represent CD8+ T cells derived from peripheral blood co-stained with IFN-γ and CD11a, following ex vivo restimulation with SIINFEKL. Data information: Data are representative of two experiments performed with scatter plots showing mean values (±SEM). (D) Protective efficacy as measured by quantitative real-time PCR. Groups of mice were vaccinated as described and challenged 19 days later with 10,000 WT (n=7 non-vaccinated, n=9 vaccinated), CSPSIINFEKL (n=6 non-vaccinated, n=4 vaccinated) or UIS4SIINFEKL sporozoites (n=3 non-vaccinated, n=11 vaccinated). 42 hours later livers were removed, and parasite load was assessed by qPCR. Plots show the relative parasite load of mice in each condition (**, p<0.01; Mann-Whitney U test). See Appendix Table S4 for exact p-values. Data information: Data are representative of two experiments performed with scatter plots showing mean values (±SEM). (E) Proportion of sterile protection after immunisation. Mice (vaccinated group, n=8; control group, n=4) were vaccinated as described and were challenged with 800 WT, CSPSIINFEKL or UIS4SIINFEKL sporozoites. Daily blood smears were taken from day 3-14 post challenge to check for parasitaemia. Data information: Data are representative of two experiments performed with scatter plots showing mean values (±SEM).
Figure 7A) Binding of TDP-43 to GU-rich RNA promotes liquid properties of TDP-43 condensates, as shown by the fusion of small condensates into larger droplets. This fluidity is impaired by the loss of RNA binding affinity or disease-linked mutations that alter phase separation, such as M337V and A321G, which could explain increased TDP-43 aggregation. B) RNA molecules that bind multiple TDP-43 molecules (orange lines) provide a scaffold and increase multivalent interactions, including N-and C-terminal domains, creating a meshwork that results in phase separation. In contrast non-specific RNA (blue lines) fails to sufficiently concentrate TDP-43 to generate the multivalent network necessary for condensation. Scale bars, 10 μm.
Figure 1Immunohistochemistry performed on 12 month-old THY-Tau22 mice showing AT100-positive neurons (green), CBP-positive nuclei (red) and DAPI-stained nuclei (blue) in the CA1 region of the dorsal hippocampus. A representative image is shown with a focus. Scale bars: 40µm and 20µm as noted. (n=5 mice). Arrows depict neurons bearing neurofibrillary tangles and do not display CBP immunostaining. The star depicts a ghost tangle (AT100-positive neuron and no nucleus) Western blot analyses perfomed in the dorsal hippocampus of 12 month-old THY-Tau22 mice compared to age-matched controls. NeuN, actin, tubulin and total H2B levels are not changed. Phosphorylated tau on serine 404 (Tau-Ph404) attest for samples from tauopathic mice. Quantification represents the ration of the protein level detected on the total amount of proteins on the membranes, with WT arbitrarily set at 1 (fold induction). Bar graphs are mean ± SEM. n=5-7/group as noted, multiple t tests, CBP * p=0.003 and GFAP **p=0.0001 for THY-Tau vs. WT mice RNA-sequencing data performed in the hippocampus of 8 month-old mice. Functional analyses (DAVID GOTERM) performed on significantly deregulated genes in THY-Tau22 mice compared to WT mice. Representative genes are presented below. |log2 Fold Change| > 0.2; * indicates FDR<0.05. IEG, Immediate Early Genes. Predicted promoter motif (right) analyses performed with GREAT. Groups: WT mice (n=3), THY-Tau22 mice (n=2) Long-term spatial memory testing: Mice were injected 3 times (1 per week) with Vehicle (WT mice saline, WT VEH, n=17), Vehicle (THY-Tau22 mice CSP, 500 μg /mouse, TAU VEH, n=10) or Molecule (THY-Tau22 mice CSP-TTK21, 500 μg/mice TAU MOL, n=13) before training of spatial memory in the Morris water maze (MWM); retention (Probe test) was tested 10 days after the last training session. Acquisition (Escape latencies, seconds) and retention performances (Time in target quadrant, seconds) are shown for the 3 groups of mice. All groups of mice displayed significant acquisition of the platform location (Day effect, F(4,148)= 26.45, p<0.001; Group and Group X Day effects, ns), but only TAU VEH exhibited impaired retention. CSP-TTK21 treatment fully restored the ability of THY-Tau22 mice to remember the platform location. Bar graphs are mean ± SEM. Student's t test to a constant value, # when compared to random (dotted line, 15sec) WT VEH, t(16)=4.6323, #p=0.0002; TAU VEH, t(9)=0.3606, p=0.7267; TAU MOL, t(12)=3.945, #p=0.0019. One-way ANOVA; F(2,37)= 3.55; p= 0.03; * in the different comparisons: TAU MOL vs. TAU VEH, *p=0.0166; WT VEH vs. TAU VEH, *p= 0.040; TAU MOL vs WT VEH, non significant p=0.437). TQ, Target Quadrant; O, Other, corresponds to the mean of the 3 other quadrants
Figure 3A) Immunoblot showing the levels of phosphorylated CHK1 (Ser345) and GAPDH in the Mybl2+/+ and Mybl2Δ/Δ ESCs with or without CPT treatment (2.5µM CPT for 4 hours). B) Immunoblot showing the levels of P-CHK1 (Ser345) and CHK1 in irradiated Mybl2+/+, and Mybl2Δ/Δ ESCs with or without 3 hours inhibitor treatment: ATR (AZ20, 5µM) and ATM (Ku60019, 10µM). Cells were also exposed to 5Gy irradiation before the final hour of inhibitor treatment. Bar graph (lower panel) represents average band density of P-CHK1 in Mybl2Δ/Δ ESCs relative to CHK1 and relative to untreated cells. Mybl2+/+ from two independent repeats and Mybl2Δ/Δ ESCs from three independent repeats. Error bars represent SD. Statistical analysis was carried out using a two-tailed unpaired t-test (*p< 0.05). C) Cells were treated for 1.5 hours with 5µM AZ20, before sequential addition of IdU and CldU for 20 minutes each. Dot plot representing the effect of ATR inhibition upon replication rate in Mybl2+/+ and Mybl2Δ/Δ ESC. Statistical analysis of distributions was carried out using unpaired Mann Whitney U tests. A minimum of 240 forks were quantified for each of the genotypes and conditions shown from 2 independent repeats (**** p< 0.0001) D) Frequency of new firing origins (relative to total structures counted) from Mybl2+/+ and Mybl2Δ/Δ ESCs treated or not with ATR inhibitor AZ20. At least 500 replication structures were counted per treatment from two independent repeats. Statistical analysis using two- tailed unpaired t-test.(*p< 0.05; ns= no significant). E) Distribution and average of IdU/CldU fork ratio in Mybl2+/+ and Mybl2Δ/Δ ESCs with or without ATR inhibitor treatment. Percentage of highly unstable forks (ratio above 2) are indicated. Dotted line at 1 indicates positive ratio. Y-axis was cut to change scale and display large values. Statistical analysis was carried out using the Mann Whitney U test. At least 150 ratios were calculated per treatment group from two independent repeats (*p < 0.05; **p< 0.01; ***p< 0.001; **** p< 0.0001).
Figure 3A- HEI-OC1 cells containing a Tlr4 deletion (Tlr4-/-) were compared to HEI-OC1 non-targeting (NT) control cells and assessed for cell viability (A), AnnexinV/propidium iodide staining (B), following cisplatin treatment at the indicated concentrations (n=3 independent biological replicates). Data Information: In all panels data are presented as mean and standard deviation. Statistical comparisons to NT at the same cisplatin concentration were assessed by 2-way (A, B) *, P<.05; **, P<.0001(Bonferroni test). HEI-OC1 cells containing a Tlr4 deletion (Tlr4-/-) were compared to HEI-OC1 non-targeting (NT) control cells and assessed for ROS generation (C) following cisplatin treatment at the indicated concentrations (n=3 independent biological replicates). Data Information: In all panels data are presented as mean and standard deviation. Statistical comparisons to NT at the same cisplatin concentration were assessed by one-way (C ANOVA. P<.01; ***, P<.0001(Bonferroni test). A-D HEI-OC1 cells containing a Tlr4 deletion (Tlr4-/-) were compared to HEI-OC1 non-targeting (NT) control cells and assessed for IL-6 secretion (D) following cisplatin treatment at the indicated concentrations (n=3 independent biological replicates). Data Information: In all panels data are presented as mean and standard deviation. Statistical comparisons to NT at the same cisplatin concentration were assessed by one-way D) ANOVA. ; ****, P<.0001(Bonferroni test).
junD-deficient fibroblasts exhibit cell cycle delay, decreased cell survival and increased DNA damage.Figure 2A Bar plot showing the doubling time of wt and junD-/- fibroblasts. B Representative cell cycle distribution of asynchronous wt and junD-/- fibroblasts. C Bar plots showing the percentage of wt and junD-/- fibroblasts in G1, S and G2 phases of the cell cycle, following serum stimulation for the indicated times (hours, h), after 48h serum starvation. D Representative views of alkaline comet assays from wt and junD-/- fibroblasts either untreated or after 10Gy γ-irradiation for the indicated times (minutes, min).E Alkaline comet assays from wt and junD-/- fibroblasts at basal state. Left, Box-plots showing tail moments at basal state in wt and junD-/- fibroblasts. Right, Distribution of wt and junD-/- fibroblasts expressed as percentage of cells (%) according to categorized tail moments. At basal state, 16% of junD-/- cells exhibit tail moment higher than 5, while it is the case for only 3% of wt cells.F Alkaline comet assays from wt and junD-/- fibroblasts after 10Gy γ-irradiation. Box-plots showing tail moments at basal state in wt and junD-/- fibroblasts, following 10Gy γ-irradiation for the indicated times (minutes, min).G Distribution of wt and junD-/- fibroblasts expressed as percentage of cells (%) according to categorized tail moments, following 10Gy γ-irradiation for the indicated times (minutes, min). H Surviving fraction of wt and junD-/- fibroblasts 7 days after 2Gy or 5Gy γ-irradiation, as assessed by clonogenic assay. Surviving fraction is expressed as percentage (%) of number of colonies after irradiation normalized with respect to untreated conditions. I Surviving fraction of wt and junD-/- fibroblasts either transfected with an empty vector (+ Ctl) or H2AX-expressing (+ H2AX) vector. Surviving fraction is expressed as percentage (%) of alive cells, 72 hours post-transfection.