EMBO/sd-character-level-ner huggingface dataset:
Figure 4(A) Representative western blot showing level of ATF4 in ATF4-silenced Flp-In 293 HA-IKKε cells treated with doxycycline (16 hours). (B) qRT-PCR analysis of PHGDH, PSAT1 and PSPH mRNA levels in Flp-In 293 HA-GFP or Flp-In 293 HA-IKKε cells treated with doxycycline (Dox) for 16 hours. Data are expressed as fold changes, relative to levels in non-treated Flp-In 293 HA-GFP cells and normalised to β-Actin (n=4 biological replicates) (C) Representative western blot showing levels of IKKε, IRF3, phosphorylated IRF3 (S396) and SBP enzymes in Flp-In 293 HA-GFP or Flp-In 293 HA-IKKε cells treated with doxycycline (Dox) for 16 hours. (D) qRT-PCR analysis of PHGDH, PSAT1 and PSPH mRNA levels in ATF4-silenced Flp-In 293 HA-IKKε cells treated with doxycycline (Dox) for 16 hours. Data are expressed as fold changes, relative to levels in a non-silenced, non-treated control, and normalised to β-Actin (n=5 biological replicates). (E) Representative western blot showing levels of IKKε, PHGDH, PSAT1 and PSPH in ATF4-silenced Flp-In 293 HA-IKKε cells treated with doxycycline (Dox) for 16 hours.
null embryos exhibit overt neural tube closure and limb-bud outgrowth defects, reduced Fgf8 expression domains and aberrant neuronal and neural crest differentiation (A-H) Live wildtype littermate and Slc7a5-null embryos imaged shortly after dissection.Figure 2(A-H) Live wildtype littermate and Slc7a5-null embryos imaged shortly after dissection. (A-D) Wildtype and (E-H) Slc7a5-null E9.75 embryos from lateral (A, D, E, H), frontal (B, F) or dorsal (C, G) views; (e1-h1) higher magnification images of the Slc7a5-null embryo. White arrowheads indicate the smaller limb bud (compare B and F), open/reduced forebrain (compare D and H), open neural tube at posterior (compare C and E, e1) and anterior (compare B and F, f1) regions. Black arrowheads indicate kinked neural tube (compare C and G, g1) and apparently missing optic and smaller otic vesicles (ov) (compare D and h1). Data information: Scale bars 200 µm mRNA in situ hybridization and immunofluorescence in E9.5 or E10.5 wildtype and Slc7a5-null embryos for key marker genes. (I-L) Fgf8 mRNA transcripts were detected in wildtype (I-j1) and in Slc7a5-null (K-l1) E10.5 embryos (n=4 each) (white arrowheads indicate limb buds in K and the isthmus in L) with (j1, l1) sections through isthmus at midbrain/hindbrain border and (i1, k1) lateral views of forelimb buds. Data information: Scale bars 200 µm, except sections 100 µm. mRNA in situ hybridization and immunofluorescence in E9.5 or E10.5 wildtype and Slc7a5-null embryos for key marker genes. (M-N) Wholemount Tubulin-β-III (Tuj1) immunofluorescence performed on (M, m1) wildtype and (N, n1) Slc7a5-null E9.5-E10.5 embryos (n=5 each condition). Dorsal root ganglia (DRG) and Sympathetic Chain Ganglia (SCG) are indicated with open arrowheads together with cranial ganglia IX, X and XI. Images in (m1, n1) show higher magnification of the cranial ganglia V, VII/VIII (open arrowheads). Data information: Scale bars 200 µm mRNA in situ hybridization and immunofluorescence in E9.5 or E10.5 wildtype and Slc7a5-null embryos for key marker genes. (O-P) Neurog2 expression in wildtype (O-o4) and Slc7a5-null embryos (P-p4) (n= 5 and n=6 each), showing reduced expression in forebrain (o2, p2), cranial ganglia (o1, o3, p1, p3, arrowheads) and in spinal cord at level of forelimb (o4, p4, arrowheads indicate position of neural crest). Data information: Scale bars 200 µm, except sections 100 µm.
Figure 7A, B hTERT-RPE1 cells transfected with non-targeting siRNA control (Neg) or RAB35 siRNA, were serum-starved for 48 h and stained for INPP5E, ARL13B, acetylated tubulin (acetyl. tub.) and DNA. Representative images are shown in (A). Regions within white boxes shown at higher magnifications to the right. Scale bars, 10 µm. (B) Box-and-whisker plots show quantification of ciliary INPP5E intensity in arbitrary units (a. u.). Horizontal lines show 25, 50 and 75th percentiles; whiskers extend to minimum and maximum values. One representative experiment out of three is shown (n > 50 cilia per experimental condition). Statistical significance according to Kruskal-Wallis followed by Dunn's post-hoc test (*** P < 0.001; P-values: Neg vs. RAB35-1 P < 0.0001, Neg vs. RAB35-2 P = 0.0002). C For co-depletion experiments, hTERT-RPE1 cells were first transfected with Neg or RAB35 siRNA. After 24h, a second transfection with Neg or ARL13B siRNA was performed and cells were serum starved for 48 h and stained as in (A). Horizontal lines show 25, 50 and 75th percentiles; whiskers extend to minimum and maximum values. One representative experiment out of three is shown (n > 40 cilia per experimental condition). Statistical significance according to Kruskal-Wallis followed by Dunn's post-hoc test (** P = 0.0017). D, E NIH3T3 wild type (WT) and Rab35 knockout (KO) cell lines were serum-starved for 24h and stained for Innpp5e, acetylated tubulin (acetyl. tub.) and DNA. Representative images are shown in (D). Higher magnifications of regions within white boxes shown at the bottom. Scale bars; 10 µm. (E) Box-and-whisker plots show quantification of ciliary Inpp5e intensity in arbitrary units (a. u.). Horizontal lines show 25, 50 and 75th percentiles; whiskers extend to minimum and maximum values. One representative experiment out of three is shown (n > 25 cilia per experimental condition). Statistical significance according to Kruskal-Wallis followed by Dunn's post-hoc test (* P < 0.05, *** P < 0.001; NIH3T3 WT vs. Rab35 KO#1 P < 0.0001, NIH3T3 WT vs. Rab35 KO#2 P = 0.0345). F, G Localisation of the PI(4,5) P2 sensor PH-PLCδ1-GFP to cilia in live NIH3T3 WT and Rab35 KO cells. Cilia are marked with HTR6-RFP. Representative images are shown in (F). Higher magnification images of the cilia region shown to the right. Scale bars; 10 µm. Quantification of localisation in (G). Data are mean ± S.E.M. of 3 independent experiments (n > 100 cilia per experimental condition). Statistical significance according to two-way ANOVA (* P = 0.0242).
Effect of TMPRSS2 and IFITM on S- or ACE2-expressing cells.Figure 4A. S-expressing cells (Donor cells), ACE2-expressing cells (Acceptor cells) were co-transfected with TMPRSS2, IFITM or control plasmids. Cell fusion was quantified by measuring the GFP+ area after 18h. The indicated combinations were tested. The impact of IFITMs was measured in absence of TMPRSS2 (left panel), in presence of TMPRSS2 in donor (middle panel) or acceptor cells (right panel). Data are mean±sd of 4 independent experiments. Statistical analysis: One-Way ANOVA, ns: non-significant, ∗∗p < 0.01, ∗∗∗∗p < 0.0001. Impact of TMPRSS2 on IFITMs levels. B. TMPRSS2 does not decrease IFITM amounts measured by flow cytometry. 293T cells were transfected with the indicated IFITM plasmids, with or without TMPRSS2, and analyzed 18h later. C. Impact of TMPRSS2 on S and ACE2, measured by Western Blot. 293T cells were transfected with or without IFITM1, TMPRSS2, S or ACE2 plasmids, and analyzed 18h later. Impact of TMPRSS2 on S levels. D. TMPRSS2 decreases S surface levels, measured by flow cytometry. 293T cells were transfected with S plasmid, with or without TMPRSS2, and analyzed 18h later. Murine polyclonal anti-S antibodies, one serum from a convalescent COVID-19 patient (sera n°111), and two monoclonal antibodies (anti-S1 and anti-S2) were tested. Data are representative of three independent experiments.
a) Cells expressing either WT GFP-VPS35 or the D620N mutant were treated with siRNA to abolish expression of endogenous VPS35.f2a) Cells expressing either WT GFP-VPS35 or the D620N mutant were treated with siRNA to abolish expression of endogenous VPS35. Each dish of cells was lysed in either HEPES lysis buffer (H) or PBS lysis buffer (P). Following centrifugation, equal portions of each lysate were combined to generate a mixed lysate (H/P). Each lysate was incubated with anti-GFP to recover the respective GFP-tagged VPS35 protein. (b) Similar to (a), but only cells expressing GFP-VPS35 wild type were analysed, either mock treated or treated with siRNA to silence FKBP15 expression. (c) Cells stably expressing WT or D620N GFP-VPS35 were treated with indicated siRNAs, lysed and immunoprecipitated as in (a).
Inactivation of c-Jun and JunB in bulge HF-SCs is sufficient to initiate psoriasis-like development A.Figure 3Experimental timeline showing the different time points analyzed after mifespristone induction (day 5, 7, 15 and 30, green) to evaluate GFP expression in ear skin from control (CoK15-mT/mG) and mutant (DKO*K15-mT/mG) mice. Ear pictures of control, DKO*, and DKO*K15 mice after 15 days of induction show clear signs of inflammation and psoriatic-like scales in both models. Representative images of hematoxylin and eosin (H&E) staining of ear sections from control, DKO* and DKO*K15 mice. Ear thickness measurement at different time points during psoriasis-like development in control, DKO* and DKO*K15 mice. n > 5 per group. Statistical significance *p<0.05, **p<0.01, ***p<0.001 (t-student two tailed-test relative to control group). FACS quantification of CD45+ cells from ear skin of control, DKO* and DKO*K15 mice at different time points during psoriasis-like development. n > 5 per group. Statistical significance *p<0.05, **p<0.01, ***p<0.001 (t-student two tailed-test relative to controls). Quantification of IL-17A, IL-6 and LCN2 production in sera of control, DKO* and DKO*K15 mice at day 30 of psoriasis-like disease by ELISA. n > 4 per group. Statistical significance *p<0.05, **p<0.01, ***p<0.001 (t-student two tailed-test relative to controls). Quantification of total number of GFP+ epidermal cells by FACS analysis from DKO*-mT/mG and DKO*K15-mT/mG mice at different time points during psoriasis-like progression compared to Co-mT/mG mice. n = 3-6 per group and time point. Representative immunofluorescence images of ear sections from CoK15-mT/mG and DKO*K15-mT/mG mouse models after 15 days of induction. Red (Tomato), green (GFP) and blue (DAPI). Hyperplasia of Tomato+ keratinocytes in psoriatic-like DKO*K15-mT/mG mouse with clusters of GFP+ keratinocytes are derived from hair follicle K15+ GFP+ stem cells. n=3 per group. Dotted lines separate epidermis and dermis. Quantification analysis of cleaved Caspase-3 (cCas3) and Ki67 in ear skin of DKO*K15-mT/mG mice at different time points during psoriasis-like disease progression.
Figure 5A. Quantitative real-time PCR detection of mRNA levels for PEX3, PEX13, PEX14, and PEX19 in HeLa cells transfected with the indicated siRNA. ***p<0.001; two-tailed unpaired t-test. B. Quantification of Parkin-mediated TOMM20 clearance in HeLa/Parkin cells transfected with the indicated siRNA 16 h after treatment with 10 µM CCCP. Results represent mean ± SEM of triplicate samples (~100 cells analyzed per sample). Similar results were observed in three independent experiments. **p<0.01, ***p<0.001; one-way ANOVA with adjustment for multiple comparisons. C. Quantification of mtDNA clearance in HeLa/HA-Parkin cells transfected with the indicated siRNA 8 h after treatment with 2.5 μM oligomycin and 250 nM antimycin A (OA). Results represent box plots of ~300 cells analyzed per sample. Whiskers represent 5%-95% range, and each outlier is represented by a dot. Similar results were observed in three independent experiments. ****p<0.0001, NS= not significant; Kruskal-Wallis H-test. D. Quantification of GFP-LC3 puncta in HeLa/GFP-LC3 cells treated with 10 nM Baf A1 or DMSO vehicle and cultured in normal medium or 3 h HBSS starvation media. Results represent mean ± SEM in triplicate samples (~100 cells analyzed per sample). Similar results were observed in three independent experiments. *p<0.05, **p<0.01, ***p<0.001, NS= not significant; one-way ANOVA with adjustment for multiple comparisons. Statistical analyses refer to the differences between Pex or ATG7 siRNAs vs. NC siRNA within each treatment group.
Figure 2(A) Primary hippocampal neurons (DIV6+4) were transduced with shRNA targeting TDP-43 or a control together with tagRFP (not depicted). Representative images of neurons subjected to Alexa-transferrin (Alexa-Tf) for 20 min (pulse) and chased for 0, 20 or 60 min in complete media. Scale bar represents 100 µm. (B) Quantification of labeled cellular Alexa-Tf normalized to cell area (using tagRFP) after chase period. At least ten images per condition were analyzed per experiment in three independent experiments. (C) ELISA measurement of β2-transferrin in CSF of control subjects (n=65) and sporadic ALS patients (n=55), FTD-ALS patients (n=12) and behavioral variant FTD patients (bvFTD, n=16). (D) Primary hippocampal neurons (DIV6+3) were transfected shTDP or shCtrl and GFP as transfection marker. After 3 days neurons were subjected to Alexa-Tf for 20 min and intracellular Alexa-Tf movement was imaged as in Figure 1 afterwards. Quantitative analysis of Alexa-Tf movement (E) and vesicle number (F) from kymographs.
Fig.Figure 5De-mannosylation (i.e., removal of light green mannoses by ER α-mannosidase I (αMI) and EDEM proteins) generates ligand for mannose-binding lectins (OS-9 and XTP-3B). These shuttle misfolded proteins to translocation sites. Translocons are built around ER membrane-embedded E3 ligases (e.g., HRD1, TEB4, GP78) and include co-factors that confer client-specificity and promote cytosolic delivery. Misfolded proteins are de-glycosylated, poly-ubiquitylated and eventually degraded by 26S-proteasomes (ERAD, upper part) (Sun & Brodsky, 2019). De/re-glucosylation cycles and persistent association with Calnexin (CNX) select ERAD-resistant misfolded proteins for segregation in ER subdomains. ER vesiculation is directly or indirectly promoted by ER-phagy receptors (e.g., FAM134B, SEC62, RTN3L, CCPG1). ER-derived vesicles containing misfolded polypeptides are delivered to the endolysosomal district by LC3-dependent transport, macro- or micro- ER-phagy (ERLAD, lower part) (Molinari, 2021).
Model for the Role of Autophagy in Lifespan Extension by Dietary Restriction in C.Figure 7In response to dietary restriction using the eat-2 mutation, TOR and RAB-10 activities fall, which triggers autophagy. Autophagy requires altered gene expression, since its appearance requires the transcription factor PHA-4. Autophagy is necessary but not sufficient for lifespan extension in long-lived daf-2 insulin/IGF-1-receptor mutants; instead, autophagy and the DAF-16/FOXO transcription factor are both required, independently, for lifespan extension. Inhibiting protein synthesis in well-fed animals may activate a distinct longevity pathway, since inhibiting protein synthesis in the context of dietary restriction produces a lifespan extension that requires autophagy gene function, whereas inhibiting protein synthesis in well-fed animals produces a lifespan extension that is independent of autophagy gene function. The sir-2 histone deacetylase is not in this diagram, as we [6] and several other groups [66-68] have found that sir-2 deletion mutations do not prevent dietary restriction from increasing lifespan in C. elegans.
Simplified metabolic networks of ASPC-1 cells as determined by proton NMR spectroscopy.Figure 6  The ASPC-1 cell line is shown without and upon various treatment regimens (OMP, 5-FU or 5-FU+OMP combination). The nodes of this network symbolize metabolite signals, their colours correspond to relative signal intensity (when compared to an external standard) as indicated in the heatmap scale below. The signal intensity is linearly related to the intracellular concentration. The background of the nodes are left blank when the signal intensity is out of the range indicated by the heatmap scale. The lines between the boxes symbolize strongly simplified metabolic pathways. The colours of these lines indicate significant differences of the signal intensity ratios of the connected metabolites compared to the control group when orange (p<0.05), red lines indicate p<0.01. The most obvious changes is that the PC/Cho ratio is significantly lower in the OMP group compared to the control group. Upon 5-FU, the Cho/Acetate ratio decrease is the only significant change. In the 5-FU+OMP group, there are several significant changes, i.e.the FACH2/CH = CH ratio is significantly higher. Moreover, the Cho/Acetate ratio changed upon 5-FU+OMP as in the 5-FU group, but also the citrate/GSH ratio. The cellular biochemical effects involve mainly the fatty acid and phospholipid metabolism pointing to membrane anabolism. Abbreviations: Gln - glutamine, Ala - alanine, PC - phosphatidylcholine, Cho - Choline, Lac1+FACH2 - methyl group signal of lactate and methylene groups of the fatty acids, Lac2 - methylene group of lactate, CH = CH - protons of methin groups of unsaturated fatty acids.
Figure 5(a) H2AX Western blot in protein extracts from control and Huwe1 deficient intestinal epithelial cells. Band intensity relative to βactin displayed under each lane (Mann-Whitney, p = 0.04, n = 3 vs 3). (b) γ-H2AX IHC showing increased positivity in Huwe1 deleted intestinal cells. Scale bars = 50 µm. (c) γ-H2AX Western blot in protein extracts from control and Huwe1 deficient intestinal epithelial cells (Mann-Whitney, p = 0.04, n = 3 vs 3). (d) γ-H2AX IHC of Vil Apc and Vil Apc Huwe1hom tumours, note increased positivity in Huwe1 deficient tumours. Scale bars = 50 µm.
a, Homology between Apg3, Apg10 and their homologues.figf3b, Interaction between Apg8 and Apg3. Lysates of Δapg3 cells expressing APG3 and/or Myc-APG8 from 2µ plasmids were immunoprecipitated with anti-Apg3 antibody, and immunoblotted with anti-Apg3 and anti-Myc antibody. c, Effect of mutated Cys 234 in Apg3 on the interaction between Apg8 and Apg3. Lysates of Δapg3 cells expressing Myc-APG8 and either wild-type (wt) APG3 or mutant APG3 (C234S and C234A) from 2µ plasmids were immunoprecipitated with anti-Myc antibody, and immunoblotted with anti-Myc and anti-Apg3 antibodies. d, Thioester formation between Apg8 and Apg3. Δapg3, Δapg7 and Δapg10 cells were transformed with both Myc-APG8 and Apg3, or only Apg3, on 2µ plasmids. Cell extracts were immunoprecipitated with anti-Myc antibody, and subjected to immunoblot (IB) using anti-Myc and anti-Apg3 antibody. DTT-, no DTT; DTT+, 100 mM DTT; asterisk, Apg8-Apg3 conjugate; diamond, possible Myc-Apg8 dimer.
Figure 2(A-C) Cohorts of animals were trained for specific parcour settings and then examined in repeated runs. Setting of hurdles was changed between the runs. Time and snout contacts were monitored. Analysis included all runs, except those in which the experimental pig did not meet the compliance criteria. Data points are plotted, lines indicate mean values (MV) ± standard deviation (SD), numbers give MV ± standard error (SE) in addition. In a barrier course (A), different arrangements of shields (see Figure S4A) were used to challenge orientation. Only minor differences between WT (blue) and USH1C (orange) pigs were observed in the dark or under normal light conditions. Data points are summarized for two separate groups at 5-12 months of age (1 USH1C, 1WT in 25 runs in the dark and 16 runs under light; 1 USH1C, 6WT in 16 runs in the dark and 11 runs under light). (B) In a similar approach, different obstacles were placed instead of shields (see Figure S5) and revealed intensified sensing by snout contacts for USH1C pigs in the dark and under light. In the light, WT pigs were also faster. 5 USH1C and 5 age-matched WT control pigs were examined in 22 runs under light conditions and 6 runs in the dark at 7-12 months of age. Statistical evaluation was performed by two-tailed Mann-Whitney U test (p-values indicated). For analysis of normalized data sets see Figure S6. (C) Detached analysis of the tests in the obstacle course (B) revealed that both, hurdles that primarily challenge the vestibular system (red cavaletti, blue F) as well as hurdles daring vision (sideboard, hanging or standing ton) provoked significantly higher alternative orientation by snout contacts for USH1C pigs. (D) Plotting individual runs from the obstacle course (B) to a planar matrix facilitated parametrizing the positions of the left (red) and right front leg (red) as well as the time when animals touch the ground (in seconds) and comparison of the trajectory to the positions of the hurdles. Data were then introduced in TrajR for evaluating standard motion parameters. Coordinates in the parcour were indicated in cm and Y-axis of the respective parameters are given according the TrajR output. Error bars and numbers indicate mean values (MV) ± standard error (SE).
Figure 2A Distribution of RAD51NES in the SCOTROC4 cohort. Dashed line denotes the median RAD51NES in this cohort. B Survival analysis of the SCOTROC4 cohort. Kaplan-Meier plots for PFS (left) and OS (right) stratified according to quartiles of RAD51NES. Log-rank test, shading denotes 95% confidence intervals. C Number of cases with progression at 12 and 24 months. Chi-square test. D Survival analysis of HRD positive patients according to quartile of RAD51NES. Log-rank test, shading denotes 95% confidence intervals. E, Survival analysis of HRD negative patients. Log-rank test, shading denotes 95% confidence intervals.
WDR18 and TEX10 co‐immunoprecipitate with PELP1.Figure 2(A, B) Flag‐PELP1 and HA‐WDR18 were expressed in HeLa cells as indicated. HA‐WDR18 was captured on HA‐beads and the bound material was probed by immunoblotting for the presence of ectopically expressed Flag‐tagged PELP1 or endogenous PELP1 as indicated. (C) Endogenous PELP1 was immunoprecipitated from HeLa cells with a rabbit polyclonal antibody and immunocomplexes were probed for the presence of endogenous WDR18 by western blotting with an anti‐WDR18 antibody. (D) Flag‐TEX10 was expressed with HA‐PELP1 in HeLa cells, captured on Flag‐beads and proteins were immunoblotted as indicated. (E) HA‐TEX10 was expressed, captured on HA‐beads and associated proteins were probed for the presence of endogenous WDR18 using immunoblotting. The inputs represent 2.5% of the total cell lysate. (F) Scheme of mutual interactions between PELP1, TEX10, WDR18 and SENP3.
Figure 3A. UMAP embeddings of transcriptional profiles from spontaneously differentiated embryoid bodies (EBs). Each point represents a cell colored by genetic background and shaded by biological replicate (N = 3). B. Similar to A indicating 13 major cell populations based on unsupervised clustering. Each cell is colored based on cluster. C. Proportion of cells in a given cluster from B represented by either B6 or D2 genetic background. D. Feature plot of expression gradients for indicated gene overlaid on UMAP from a. Three lineage markers that help to distinguish cellular identity are shown for five cell clusters. E. Pseudocolor plots of FACS analysis on cells from EBs for both B6 and D2 strains. Cells were labeled with anti-EpCAM (top) or biological replicates with anti-SOX1 antibody (bottom). Gated population indicates percentages of EpCAM+ (Q3) or SOX1+ cells. F. Bar chart showing percent positive cells gated for EpCAM+ population (left) or SOX1+ population (right). As predicted by scRNA-seq, more D2 EB cells express EpCAM compared to B6 and more B6 EB cells express SOX1 compared to D2 (N = 3, two-sided t-test, EpCAM p-value = 0.0012, and SOX1 p-value = 0.0048).
Figure 3A. Ribbon diagram showing the 2-fold dimerization interface of Mdm12. Oxygen and nitrogen atoms are shown in red and blue, respectively. The orange dotted lines indicate intermolecular hydrogen bonds between two protomers of Mdm12. The sequence alignment of yeastMdm12 orthologs is shown to highlight the sequence conservation in the N-terminus β1-strand. Ten orthologs are aligned from residues 1 to 11. Absolute and highly conserved residues are indicated in red and orange, respectively. B. The molecular weight of the Mdm12 (I5P) mutant was measured by size-exclusion chromatography (below) and ultracentrifugation (top) as in Figure 1B and C. C. Ribbon diagram showing the structures of the SMP domain in E-SYT2, CETP, and BPI for the comparison of dimeric interfaces among SMP domains. Note that CETP and BPI are not dimers but monomers containing two tandem SMP domains.