EMBO/sd-character-level-ner huggingface dataset:
scRNA-Seq identifies highly variable homeodomain TF expression in late embryonic MNs at the time of synaptic wiring.Figure 1(A) Schematic drawing depicts the step-wise development of the nervous system in Drosophila, starting during embryogenesis, progressing through three larval stages and metamorphosing during pupal stage into the adult nervous system. The first connections in the neuromuscular system are formed between MNs and muscles in late Drosophila embryos (stage 16). MNs at this stage expressing UAS-RFP under the control of the motoneuronal driver OK371-GAL4 (OK371>RFP) are color-coded along the ventral nerve cord according to different patterns of Hox gene expression. OK371-RFP positive MNs were FACS sorted and single cells were sequenced by targeted Smart-Seq2 to enrich for Hox gene representation (HoxSeq) as spatial markers (see also Materials and Methods). (B) t-distributed stochastic neighbor embedding (t-SNE) plot of n=999 single-cell transcriptomes. Colors correspond to three clusters, VUM neurons (purple), twitlow (grey) and twithigh (green) MNs. (C) Identification of highly variable genes in the twitlow cluster using the method by (Brennecke et al, 2013). Scatter plot depicts the mean expression for each gene and squared coefficient of variation across twitlow cells. The solid line indicates the fit, dashed lines the 95% confidence interval. Genes with a significantly elevated variance are shown as triangles, other genes as circles. Different gene classes are color coded. p-values shown were retrieved by a hypergeometric test for enrichment of the respective gene class among highly variable genes. (D) Outline of strategy to map single MNs to a position along the AP axis (see also Materials and Methods). Upper panel: Intensities of Hox protein expression along the ventral nerve cord measured by immunofluorescence (upper left panel) and co-expression patterns of Hox gene transcripts measured by scRNA-Seq (upper right panel) were used as input. Upper right panel depicts a heatmap of color-coded Hox gene expression levels; columns correspond to n=758 single twitlow MNs. Lower panel: AP position is inferred form scRNA-Seq data by probabilistically mapping Hox gene expression pattern in each individual cell to the immunofluorescence reference data. (E) Genes with significant variation along the AP axis were identified and clustered into 10 groups of distinct expression patterns (Materials and Methods). Heatmap shows average gene expression per cluster (rows) across single cells (columns). Asterisks indicate p-value of a hypergeometric test for enrichment of protein domains, ***: p < 0.001. (F) Left panel: ZINB-WaVE (Risso et al, 2018, 2019) was used to statistically separate gene expression variability into parts linked to AP position and parts independent thereof. Scatter plot of ZINB-WaVE loadings separates known dorsal and ventral marker genes on ZINB-WaVE component 1. Right panel: Genes encoding homeodomain TFs and genes encoding Ig domain molecules (see color code) show high loadings on ZINB-WaVE component 1 and 3, demonstrating high variability independent of AP position.
A minimal 46 aa region is required for Nup50 NPC localization (A) HeLa cells were transfected with EGFP-Nup50 and different truncations.Figure 5(A) HeLa cells were transfected with EGFP-Nup50 and different truncations. After 24h cells were shortly pre-treated with 0.1% Triton X-100 in PBS, fixed with 4% PFA and analyzed by confocal microscopy (left panel). Scale Bar: 10 µm. The graph (right panel) shows the Nup50 domains (blue: Importin α interaction domain, green: NPC targeting domain and pink: Ran binding domain) and the truncation tested. Ability to bind the nuclear rim is indicated on the right. (B) Sequence alignment of Nup50 144-189, species are indicated on the left, the black boxes highlight residues tested by single point mutation in (C) and (D), and the residues number are shown above or below the alignment. The color scheme indicates the type of amino acids according to the alignment software default setting. (C) HeLa cells were transfected with EGFP-Nup50 comprising different point mutations in the minimal NPC binding region and analyzed as in (A). Scale Bar: 10 µm. (D) GST-fusion constructs of the Xenopus Nup50 minimal NPC binding fragment (aa 144-189) comprising no or single point mutations were incubated with Xenopus egg extracts. Starting material as well as bound proteins were analyzed by Western blotting with indicated antibodies. The quantitation shows the average MEL28/ELYS and Nup153 bead bound signal from three independent experiments, normalized to the signal of their respective wild-type GST fusion Data points from individual experiments are indicated.
Apelin inactivation reduces sunitinib therapy-induced metastasis and stratifies sunitinib therapy responses in patients.Figure 6(A) Number of metastatic lung foci in untreated (control) and sunitinib-treated (60mg/kg, three times a week from tumor initiation) NeuT;Apln+/+ or NeuT;Apln-/- mice, assessed 6 weeks after mammary tumors were size-matched. Data of individual lung sections and means (black bars) are shown. Right panels show representative H&E images, where black arrows and insets indicate metastatic foci. Scale bars = 1000 μm (large panels) and 50 μm (insets). *P<0.05, **P<0.01; ***P<0.001; Kruskal-Wallis test; n=3 mice per cohort and three sections per lung were analysed. (B) Kaplan Meier survival plot for distant metastasis free survival from the KM-plotter database (Data Ref: Győrffy et al, 2013) for high and low Apelin (APLN)-expressing groups in human breast cancer. Patients were split by the median. Affymetrix Apelin ID 222856_at. (C) Kaplan-Meier plot for progression-free survival stratifying RCC patients with high and low APELIN serum levels 3-5 months after the start date of sunitinib treatment. *P=0.0367; Log rank test. (D) Kaplan Meier plots for progression-free survival in RCC patients stratified into groups of high or low levels of APELIN and VEGF. Cut-off levels were set by the median. Serum was analysed 3-5 months after the start date of sunitinib treatment. *P<0.05, **P<0.01; Log rank test.
Figure 2(A) qPCR measurement of IL-1β, and TNF-α mRNA levels in WT and Irgm2-/- BMMs following 8-hour stimulation with LPS (1 µg/ml). (B) WT, Irgm2-/- and Casp1-/-Casp11-/- BMMs were treated for 24 hours with LPS at indicated doses and supernatant TNFα, IL-1β, and IL-18 was measured by ELISA. (C) IFNγ-primed WT and Irgm2-/- BMMs were treated with LPS, Pam3CSK4, poly(I:C) or a combination of Pam3CSK4 and poly(I:C) (1 µg/ml for all treatments) for 24 hours and cell supernatant IL-1β and IL-18 concentrations were assessed by ELISA. (D) WT, Irgm2-/-, Casp1-/-Casp11-/- and Irgm2-/-Casp1-/-Casp11-/- BMMs were treated with LPS (1 µg/ml) and IL-1β, IL-18, and LDH release were assessed at 24 hours post treatment (hpt). (E) IFNγ-primed WT, Irgm2-/-, Casp1-/-Casp11-/- and Irgm2-/-Casp1-/-Casp11-/- BMMs were treated with LPS (5 µg/ml) for 4 hours and subsequently stained with anti-ASC antibody and Hoechst stain (DNA/nuclei). Representative images are shown with white arrows pointing at ASC specks. Number of ASC specks per nuclei was quantified. Scale bars: 20 µm. (F) IFNγ-primed WT, Irgm2-/-, Casp1-/-Casp11-/- and Irgm2-/-Casp1-/-Casp11-/- BMMs were treated with LPS (1 µg/ml) for 24 h and cell lysates and supernatants collected. Protein levels in cell lysates (caspase-1, and actin) and supernatants (caspase-1 p20) were visualized via immunoblotting.
Figure 11(A-D) Numbers of offspring from crosses of hAPP-J20 mice with hTau-A152T(L1) (A, B) or hTau-WT(L32) (C, D) mice that were (A, C) or were not (B, D) treated with DOX. (E) Numbers of offspring from crosses of lower expresser hAPP-J9 mice with hTau-A152T(L1) mice that were not treated with DOX. n = 141 (A), 112 (B), 94 (C), 83 (D), and 282 (E) mice per cohort. Yields from two crosses indicated significant deviations from Mendelian inheritance: hAPP-J20 x TRE-hTau-A152T(L1) x CaMKII-tTA without DOX (Holm-adjusted P = 0.003, unadjusted P = 0.0008) and hAPP-J9 x TRE-hTau-WT(L32) x CaMKII-tTA without DOX (adjusted and unadjusted P < 0.0001) by chi-square goodness-of-fit test. (F) Subdural EEG recordings from untreated 3-10-month-old mice were analyzed to compare their number of spikes at baseline. n = 8 NTG, 6 hAPP-J9, 6 hTau-A152T(L1), and 8 hAPP-J9/hTau-A152T(L1). *P < 0.05, **P < 0.01 by logistic regression analysis for three-way interaction of the three transgenes with Holm adjustment (A-E) or by two-tailed Welch's t test with Holm adjustment (F).
Fig.Figure 2(A) Western blot assay of SOX1 in control and SOX1KO cells on day 16 of dorsal neural differentiation. (B) Immunostaining assay of TBR2 (red) in control and SOX1KO cells on day 25 of dorsal neural differentiation. Scale bar, 100 μm. (C and D) Immunostaining assay of MAP2 (green) in control and SOX1KO cells on day 35 of dorsal neural differentiation (C), and quantiﬁcation of MAP2+ cells (D), p=0.0001. Scale bar, 100 μm. Data information: Data are presented as mean ±SEM from three independent experiments (n=3). ***p<0.001 versus Ctrl group (unpaired two-tailed Student's t-test). (E) Immunostaining assay of NKX2-1 (red) in control and SOX1KO cells on day 25 of ventral neural differentiation. Scale bar, 100 μm. (F and G) Immunostaining assay of GABA (green) in control and SOX1KO cells on day 35 of ventral neural differentiation (F), and quantiﬁcation of GABA+ cells (G), p=0.0001. Scale bar, 100 μm. Data information: Data are presented as mean ±SEM from three independent experiments (n=3). ***p<0.001 versus Ctrl group (unpaired two-tailed Student's t-test). (H and I) Immunostaining assay of MAP2 (green) after SOX1 overexpression in V1-PAKI cells on day 35 of dorsal neural differentiation (H), and quantiﬁcation of MAP2+ cells (I), p=0.0001 (V1-PAKI), p=0.0001 (V1-PAKI+ptSOX1+dox). Scale bar, 100 μm. Data information: Data are presented as mean ±SEM from three independent experiments (n=3). ***p<0.001 versus Ctrl group ; ###p<0.001 versus -dox group (one-way ANOVA). (J and K) Immunostaining assay of GABA (green) after SOX1 overexpression in V1-PAKI cells on day 35 of ventral neural differentiation (J), and quantiﬁcation of GABA+ cells (K), p=0.0001 (V1-PAKI), p=0.0002 (V1-PAKI+ptSOX1+dox). Scale bar, 100 μm. Data information: Data are presented as mean ±SEM from three independent experiments (n=3). ***p<0.001 versus Ctrl group ; ###p<0.001 versus -dox group (one-way ANOVA).
Palbociclib blocks paclitaxel induced mitotic defects and apoptosis in the human hair follicle matrixA) Schematic of experimental design.Figure 5A) Schematic of experimental design. Hair follicles (HFs) were pre-incubated with the CDK4/6 inhibitor palbociclib for 18h, followed by a further incubation period with and without paclitaxel (or paclitaxel alone) for an additional 24h. B) Palbociclib-only and palbociclib plus paclitaxel dual treated HFs show marked reductions in Ki-67 expression in the hair matrix beyond that seen with just 24h treatment Data also confirms that Ki-67 expression is unaffected by paclitaxel treatment Analysis performed using N of 6-9 HFs per condition from 3 patients. Ordinary one-way ANOVA with multiple comparisons performed. Adjusted p values = 0.0001[***] and 0.0001 [****] respectively. Values plotted represent the mean number of positive cells counted per HF analysed. Error bars are standard error. C) Data confirming that paclitaxel treatment significantly increases (adjusted p value = 0.0001[***]) the number of pH3+ cells in the hair matrix This effect was not observed when paclitaxel treated hair follicles were pre- and co-incubated with palbociclib. Analysis performed using N of 6-9 HFs per condition from 3 patients Values plotted represent the mean number of positive cells counted per HF analysed. Error bars are standard error. D) Cleaved caspase 3 analysis in hair follicles from a patient donor that showed apoptotic sensitivity to paclitaxel treatment (adjusted p value = 0.0056 [**]). Paclitaxel treated hair follicles pre- and co-incubated with palbociclib do not show enhanced cleaved caspase 3 expression, contrasting with paclitaxel-only treatment. Ordinary one-way ANOVA with multiple comparisons performed using N of 4-5 HFs per condition. Values plotted represent the mean number of positive cells counted per HF analysed. Error bars are standard error. E) Immunofluorescence data representing how palbociclib and/or paclitaxel affect (i) Ki-67 expression, (ii) pH3 immunoreactivity and (iii) cleaved caspase 3 expression. 20 µm scale.
a) Chemical structure of SAR405.figf1(a) Chemical structure of SAR405. (b) SAR405 inhibits the function of Vps34 in the GFP-FYVE cellular assay. HeLa cells stably transfected with GFP-FYVE were incubated with DMSO (left) or with 1 μM SAR405 (right) for 2 h. Cells were fixed, and nuclei were stained using Hoechst 33342 (blue). Fluorescence was analyzed using an imaging cytometer (X40). One representative image out of three independent experiments is depicted. Scale bars, 20 μm. (c) SAR405 does not prevent phosphorylation of Akt and S6. U87MG cells were treated with SAR405 or PI103 for 2 h at the indicated concentrations. The effect of compounds on phosphorylated Akt (p-Akt) Ser473 and Thr308 residues, total Akt and on phosphorylated S6 (Ser240 and Ser244 residues) and total S6 protein was measured using western blotting. One representative experiment from at least two independent experiments is shown. Full-length blots are shown in Supplementary Figure 6.
TP53INP1-deficient mice have moderate redox-related insulin resistance syndrome which is exacerbated by HFD protocol.Figure 2A, B Histograms show bloodglucose (A) or plasmainsulin (B) levels of 6-h-fasted mice at the beginning (Week 0) and/or at the end of the protocol (Week 16). Fasting bloodglucose week 16: P (−/− versus +/+; HFD) = 0.0052; P (CTRL versus HFD; −/−) = 0.000081; P (NAC versus no NAC; −/− HFD) = 0.0019; P (w16 versus w0; +/+ CTRL) = 0.012; P (w16 versus w0; −/− CTRL) = 0.050; P (w16 versus w0; −/− HFD) = 0.0023. Fasting plasmainsulin: P (−/− versus +/+; HFD) = 0.043; P (CTRL versus HFD; +/+) = 0.028; P (CTRL versus HFD; −/−) = 0.013; P (CTRL versus HFD; +/+ NAC) = 0.011; P (CTRL versus HFD; −/− NAC) = 0.0015; P (NAC versus no NAC; −/− HFD) = 0.038. C Glucose tolerance test (GTT) was performed on 6-h-fasted mice during 120 min after injection of 1 g glucose/kg of body weight. Curves on the left show blood glucose level monitored after injection of glucose. Histograms on the right show area under curve (AUC). Fasting blood glucose: P (−/− versus +/+; CTRL; t = 0 min) = 0.045; P (−/− versus +/+; CTRL; t = 15 min) = 0.013; P (−/− versus +/+; CTRL; t = 30 min) = 0.016; P (−/− versus +/+; CTRL; t = 60 min) = 0.030; P (−/− versus +/+; CTRL; t = 90 min) = 0.017; P (−/− versus +/+; HFD; t = 60 min) = 0.041; P (−/− versus +/+; HFD; t = 90 min) = 0.043; P (−/− versus +/+; HFD; t = 120 min) = 0.034; P (HFD versus CTRL; +/+; t = 15 min) = 0.0076; P (HFD versus CTRL; +/+; t = 30 min) = 0.0067; P (HFD versus CTRL; +/+; t = 60 min) = 0.00058; P (HFD versus CTRL; +/+; t = 90 min) = 0.0010; P (HFD versus CTRL; +/+; t = 120 min) = 0.023; P (HFD versus CTRL; −/−; t = 60 min) = 0.032. AUC: P (−/− versus +/+; CTRL) = 0.023; P (−/− versus +/+; HFD) = 0.035; P (HFD versus CTRL; +/+) = 0.042.D Insulin tolerance test (ITT) was performed on 6-h-fasted mice during 150 min after injection of 0.70 U insulin/kg of body weight. Curves on the left show blood glucose level monitored after injection of insulin. Histograms on the right show area above curve (AAC). Fasting blood glucose: P (−/− versus +/+; CTRL; t = 15 min = 0.012; P (−/− versus +/+; CTRL; t = 30 min) = 0.022; P (−/− versus +/+; HFD; t = 0 min) = 0.027; P (−/− versus +/+; HFD; t = 15 min) = 0.011; P (−/− versus +/+; HFD; t = 30 min) = 0.0037; P (−/− versus +/+; HFD; t = 60 min) = 0.0028; P (−/− versus +/+; HFD; t = 90 min) = 0.041; P (−/− versus +/+; HFD; t = 120 min) = 0.0032; P (−/− versus +/+; HFD; t = 150 min) = 0.0025; P (HFD versus CTRL; +/+; t = 15 min) = 0.0082; P (HFD versus CTRL; +/+; t = 30 min) = 0.033; P (HFD versus CTRL; +/+; t = 90 min) = 0.047; P (HFD versus CTRL; +/+; t = 150 min) = 0.028; P (HFD versus CTRL; −/−; t = 15 min) = 0.026; P (HFD versus CTRL; −/−; t = 30 min) = 0.0095; P (HFD versus CTRL; −/−; t = 60 min) = 0.0031; P (HFD versus CTRL; −/−; t = 90 min) = 0.033; P (HFD versus CTRL; −/−; t = 120 min) = 0.0068; P (HFD versus CTRL; −/−; t = 150 min) = 0.0082. AAC: P (−/− versus +/+; HFD) = 0.030; P (HFD versus CTRL; −/−) = 0.037.
Figure 4(a) Top: schematic representation of the Sµ region of CH12F3-2A cells. Bottom: schematic representation of the Sµ region of CH12F3-2A cells. The 5′ Sµ region analyzed for SHM is underlined. The genomic DNA isolated from WT or Samhd1 KO CH12F3-2A cells from three independent experiments were pooled prior to SHM analysis. (b) Mutation frequency in the 5′ Sµ region of WT or Samhd1 KO CH12F3-2A cells. Statistical significance as evaluated by Fisher's exact test is shown. ns = not significant. (c) Details of the mutation analysis of the 5′ Sµ region. (d) LM-PCR analysis of the AID-induced DNA double-strand breaks in the Sµ region of WT or Samhd1 KO CH12F3-2A cells. Wedges indicate a three-fold increase in the DNA amount. Amplification of Gapdh was used as an internal loading control. (e) Gamma-H2AX-ChIP signal in the Sµ region is shown as the fraction of immunoprecipitated DNA (% IP) normalized to the total amount of DNA used for the immunoprecipitation (n=3; mean + sd; two-tailed unpaired Student's t-test; ns = not significant). (f) Left: Scheme of long-range interactions between Sμ-Sα elements in the IgH locus before and after AID activation. Right: Representative gel picture of the 3C assay detecting the Sμ-Sα interaction in WT or Samhd1 KO CH12F3-2A cells with or without CIT stimulation for 24 h. Gapdh was amplified as a loading control.
Figure 7(A) S2 cells expressing RDH/CH2064-GFP were treated with the protein synthesis inhibitor cycloheximide (CHX) for the indicated time (h: hours). The level of RDH/CH2064-GFP was determined by western blotting using anti-GFP antibody. Tubulin was used as loading control. (B) Quantification of the relative RDH/CH2064-GFP protein levels in (A). Error bars represent ± SEM. (C) S2 cells expressing Bmm-GFP were treated with the protein synthesis inhibitor cycloheximide (CHX) for the indicated time (h: hours). The level of Bmm-GFP was determined by western blotting using anti-GFP antibody. Tubulin was used as loading control. (D) Quantification of the relative Bmm-GFP protein levels in (C). Error bars represent ± SEM. (E) The RDH/CG2064-GFP protein level in 3rd instar larval fat cells is increased in CSN2 or CSN7 RNAi, as detected by anti-GFP antibody in a western blot. Tubulin was used as loading control. (F) Oil red staining of 3rd instar larval oenocytes in different genetic backgrounds under fed or starvation conditions. (G) The survival rate of control and CSN2 RNAi larvae at various fasting time points. Error bars represent ± SEM. *: P < 0.05; **: P < 0.01; ***: P < 0.001; NS: non-significant. (H) Schematic model showing how the CSN-RDH/CG2064-Plin2 axis inhibits Bmm and promotes the formation of large lipid droplets under high-fat/increased TAG conditions.
Fig.Figure 5(a) CCR9 expression in the small intestine (SI) and colon of Gallus gallus and Lonchura striata. (b) CCR9 expression in the small intestine (SI) and colon of Chrysemys picta bellii and Pelodiscus sinensis. (c) CCR9 expression in the small intestine (SI) and colon of Rana catesbiana Shaw and Rana nigromaculata. (d) GPR15 expression in the small intestine (SI) and colon of Gallus gallus and Lonchura striata. (e) GPR15 expression in the small intestine (SI) and colon of Chrysemys picta bellii and Pelodiscus sinensis. (f) C10ORF99 expression in the small intestine (SI) and colon of Gallus gallus and Lonchura striata. (g) C10ORF99 expression in the small intestine (SI) and colon of Chrysemys picta bellii and Pelodiscus sinensis. (h) GPR15 and C10ORF99 expression in the small intestine (SI) and colon of Rana catesbeiana Shaw and Rana nigromaculata. Data information: All data shown are representative of at least three independent technical replicates. Error bars indicate SD (n=3). UD, undetermined, ***P-value < 0.001; **P-value < 0.01; NS: no significance (t-test). (i) GPR15 expression in CD4+ T cell from chicken small intestine (SI) and colon. Data information: All data shown are representative of at least three independent technical replicates. Error bars indicate SD (n=3). UD, undetermined, ***P-value < 0.001; **P-value < 0.01; NS: no significance (t-test). (j) The migration of CD4+ cells from chicken small intestine (SI) and colon induced by ligand C10ORF99. Data information: All data shown are representative of at least three independent technical replicates. Error bars indicate SD (n=3). UD, undetermined, ***P-value < 0.001; **P-value < 0.01; NS: no significance (t-test).
Figure 7Diagram of the break-induced replication (BIR) assay used in this study. Red line represents galactose-inducible HO endonuclease cut site. Measurement of BIR efficiency in cells carrying an empty vector or expressing DPB11BRCT3/4 fused to wild-type or mutant versions of SGS1. Mutants include a helicase deficient allele of SGS1 (SGS1hd) or an allele lacking the Top3 Interacting Motif (TIM; SGS1TIM). Error bars represent SEM of at least 3 replicate experiments. P value was calculated with a two-tailed, unpaired t-test. * = p≤0.05; ** = p≤0.01; *** = p ≤0.001. C. Measurement of BIR efficiency in cells carrying an empty vector or expressing DPB11BRCT3/4 fused to either SGS1, TOP3, or RMI1. Error bars represent SEM of at least 3 replicate experiments. P value was calculated with a two-tailed, unpaired t-test. * = p≤0.05; ** = p≤0.01; *** = p ≤0.001. D. Measurement of BIR efficiency in cells carrying an empty vector or DPB11BRCT3/4 fused to either SGS1, hBLM, PIF1, or RRM3. Error bars represent SEM of at least 3 replicate experiments. P value was calculated with a two-tailed, unpaired t-test. * = p≤0.05; ** = p≤0.01; *** = p ≤0.001. Diagram of the gene conversion (GC) assay used in this study. Red line represents galactose-inducible HO endonuclease cut site. Measurement of gene conversion efficiency in cells carrying an empty vector or expressing DPB11BRCT3/4 fused to wild-type or mutant versions of SGS1. Mutants include a helicase deficient allele of SGS1 (SGS1hd) or an allele lacking the critical amino acid residues responsible for binding to Top3 (SGS1TIM). Error bars represent SEM of at least 3 replicate experiments. P value was calculated with a two-tailed, unpaired t-test. * = p≤0.05; ** = p≤0.01; *** = p ≤0.001. Diagram of the single-strand annealing (SSA) assay used in this study. Red line represents galactose-inducible HO endonuclease cut site. Measurement of single strand annealing in cells carrying an empty vector or expressing DPB11BRCT3/4. Error bars represent SEM of at least 3 replicate experiments. AA strain contains perfect homology on either side of a GAL-inducible break. FA strain contains 3% sequence divergence on either side of the break (homeologous product). Ratio AA / FA repair product computed from data in (H). Error bars represent SEM of at least 3 replicate experiments. P value was calculated with a two-tailed, unpaired t-test. * = p≤0.05; ** = p≤0.01; *** = p ≤0.001.