EMBO/sd-character-level-ner huggingface dataset:
Figure 5Crawling trajectories of control and neuronal TDP-43 expressed larvae as indicated. Relative crawling mobility of larvae was quantified from the indicated genotypes. Walking speed of 10-day old adult male flies of the indicated genotypes was calculated Mean ± SD. Unpaired two-tailed t-test, biological replicates: Control, n = 63, WT, n = 45, T199A, n = 70, T233A, n = 87; 2TA, n = 43. O-GlcNAc and protein levels of TDP-43 in fly head extracts were examined by immunoblotting. The asterisks in different panels indicate heavy-chain bands.    .Adult flies were fed with regular sucrose diet (RS) or high sucrose diet (HS), and survival percentage of adult flies of the indicated genotypes were examined. Kaplan-Meier survival analysis, n ≥ 40 for each group.   regular sucrose diet (RS) or high sucrose diet (HS) samples were subjected to immunoblotting and TDP-43 protein levels were examined. Walking speed of 10-day old adult male flies of the indicated genotypes was calculated. Mean ± SD. Unpaired two-tailed t-test, biological replicates: Control, n = 185, OGT, n = 148, TDP-43, n = 177, TDP-43/OGT, n = 187. Expression levels of OGT and TDP-43 were examined by immunoblotting. The asterisk represents a non-specific band. The black arrow indicates the HA (OGT) bands. Walking speed of 10-day old adult male flies of the indicated genotypes was calculated. Mean ± SD. Unpaired two-tailed t-test, biological replicates: Control, n = 186, OGT, n = 150, TDP-43 2TA, n = 207, TDP-43 2TA/OGT, n = 207. Expression levels of OGT and TDP-43 2TA mutant were examined by immunoblotting.
Figure 6(A) Left: GFP pull-down assay from whole cells extracts of BRCA2 deficient DLD1 expressing BRCA2 (WT) or the variant T207A (T207A). DLD1 BRCA2+/+ (+/+) cell extracts were used as control for the GFP-trap. DDX5 and BRCA2 were detected with specific antibodies against DDX5 and BRCA2, respectively. StainFree images of the gels before transfer was used as loading control (cropped image is shown). Right: Quantification of the GFP-trap pull-down experiments calculated as the co-immunoprecipitated DDX5 with either BRCA2 WT or BRCA2-T207A relative to the input levels of DDX5 and the amount of immunoprecipitated EGFPMBP-BRCA2. Results are presented as the fold change compared to the BRCA2 WT clone. The data represents the mean ± SD of three independent experiments. Statistical significance of the difference was calculated with unpaired t test and the p-values show the significant difference. (B) Left: Representative images of S9.6 immunofluorescence of DLD1 cells bearing BRCA2 (WT) or BRCA2-T207A (T207A). The merged images show the signal of S9.6, nucleolin (nucleoli) antibodies and DAPI staining. Scale bar indicates 25 µm. Right: Quantification of the average nuclear intensity of S9.6 antibody in DLD1 BRCA2-WT (WT) or BRCA2-T207A (T207A) cells transfected with either a plasmid expressing GFP alone (-RH) or GFP-RNase H1 (+RH), as indicated. The red line in the plot indicates the median and each symbol represents the value of a single cell. Data correspond to at least 170 cells per condition from two independent experiments. The statistical significance of the difference was calculated with Mann-Whitney U-test; p-values show the significant difference. (C) Relative DRIP-qPCR signal values (respect to the siC level at each locus) at the MALAT1, RRPH1 and HIST1H2BG loci in DLD1 BRCA2-/- cells bearing BRCA2 (WT) or BRCA2-T207A (T207A) and treated in vitro with RNase H1 (RH) pre-immunoprecipitation where indicated. The data represent the mean ± SEM from seven independent experiments. The statistical significance of the difference was calculated with paired t-Student test; the p-values show the significant difference. (D) Top: Representative images of In situ PLA performed between BRCA2 and DDX5 antibodies in cells bearing BRCA2 (WT) or BRCA2-T207A (T207A), as indicated. Cells were fixed directly or 4h post-irradiation (6Gy). Single antibody controls in non-irradiated BRCA2 WT cells are shown. Scale bar indicates 10 µm. Bottom: Quantification of the number of PLA spots per nucleus. At least 750 cells were counted per condition from three independent experiments. For statistical comparison of the differences between the samples we applied a Kruskal-Wallis test followed by Dunn's multiple comparison test and the p-values show significant differences. The red line in the plot indicates the median and each symbol represents a single PLA spot.
Figure 5(A) Chromosomal positions of various histone H3 genes, ash1 and marker transgenes. 23 histone gene repeat units, each containing single His1, His2B, His2A, His4, and His3.2 gene, are clustered near the centromere (black circle) of chromosome 2. These histone genes are removed by the ∆HisC deletion. To select the animals homozygous for the deletion, the ∆HisC chromosomes are marked with insertions of either Elav-Gal4 or UAS-2xYFP transgenes (black triangles on chromosome 2L). The His3.3A and His3.3B genes encode the same protein, but reside on chromosomes 2L and X. The transgenes carrying twelve copies of either the wild-type histone repeat unit (12xWT) or the unit in which H3 gene is altered to have K36 replaced with arginine (12xH3K36R) are inserted in the same attP site (black triangle on chromosome 3L) [47]. The ash1 gene is located on the same chromosome arm. (B) The ∆HisC; 12xH3K36R and control ∆HisC; 12xWT flies show no homeotic transformations and are indistinguishable from the wild-type.
as a ceRNA, to regulate the expression of BMP4.Figure 5A. A luciferase reporting system was used to detect the regulatory effects of gga-mir-12211 on the activity of BMP4-3'UTR (Data are shown as mean ± SEM, n =3 independent experiments, * p < 0.05, **** p < 0.0001, one-way ANOVA). B. qRT-PCR was used to detect the regulatory effects of gga-mir-12211 on LncBMP4 expression (Data are shown as mean ± SEM, n =3 independent experiments, *** p < 0.001, **** p < 0.0001, one-way ANOVA). C. The results of the luciferase reporting system showed that LncBMP4 could rescue the gga-mir-12211 inhibitory effects against BMP4-3'UTR (Data are shown as mean ± SEM, n =3 independent experiments, * p < 0.05, **** p < 0.0001, one-way ANOVA). D. Distribution diagram of RRACH sites on LncBMP4, BMP4-3'UTR, and gga-mir-12211. Enrichment levels of m6A in LncBMP4 were detected by RIP-qPCR (Data are shown as mean ± SEM, n =3 independent experiments, **** p < 0.0001, one-way ANOVA). F. Enrichment levels of m6A in BMP4-3'UTR were detected by RIP-qPCR (Data are shown as mean ± SEM, n =3 independent experiments, **** p < 0.0001, one-way ANOVA). G. The effects of MA and NPC on m6A enrichment levels in the P2 region of LncBMP4 were detected by RIP-qPCR (Data are shown as mean ± SEM, n =3 independent experiments, *** p < 0.001, **** p < 0.0001, one-way ANOVA.). H and I. The effects of m6A modifications of LncBMP4 on the ceRNA system were detected by the luciferase reporting system (Data are shown as mean ± SEM, n =3 independent experiments, * p < 0.05, **** p < 0.0001, one-way ANOVA.).
Figure 1(A) A schematic of IFITM3 protein is shown, with gray boxes indicating transmembrane domains and colored bars indicating residues targeted for post-translational modification: ubiquitinated lysines (blue), palmitoylated cysteines (purple), and a phosphorylated tyrosine (red). Residues 17-24 are shown in detail to highlight two adjacent, overlapping motifs that recruit NEDD4 (PPNY) and the AP-2 complex (YEML). The IFITM3 Δ1-21 variant initiates translation at an internal methionine residue (green) and thus lacks these motifs. (B) Confocal fluorescence microscopy of 293T cells transfected with 0.5 μg pQCXIP encoding IFITM3 variants containing N-terminal FLAG (pQCXIP-FLAG-IFITM3) and immunostained with anti-FLAG M2 antibody. Scale bar, 5 μm. (C) Flow cytometry analysis of transfected cells in (B) immunostained with anti-FLAG M2 antibody. Corresponding mean flourescence intensity values are shown. (D) SDS-PAGE of cell lysates derived from 293T cells transfected in (B) followed by immunoblotting with anti-FLAG M2 and anti-actin antibodies. (E) SDS-PAGE of Opti-prep purified supernatants (25 ng p24 equivalent) derived from 293T cells transfected with pQCXIP-FLAG-IFITM3 variants, HIV-1 pNL4-3, and Vpr-Blam plasmid followed by immunoblotting with anti-FLAG M2 and anti-p24 Gag (183-H12-5C). (F) Opti-prep purified supernatants from (E) were incubated with SupT1 target cells for 4 hours and virus-cell fusion was scored by flow cytometry. Mean + SD of 5-10 experiments (each performed with virus produced from an independent transfection) is shown. (G) Virus used in (E) were used to infect SupT1 cells and productive infection was scored at 48-60 hours by immunostaining with the KC57 antibody and flow cytometry. Expression of pCMV-MxA served as a negative control. The mean + SD of 5-10 experiments is shown. (H) Confocal fluorescence microscopy of 293T cells transfected with 0.5 μg pQCXIP-FLAG-IFITM3 variants and immunostained with anti-FLAG M2 antibody (I) SDS-PAGE of cell lysates derived from 293T cells transfected in (H) followed by immunoblotting with anti-FLAG M2 and anti-actin antibodies. (J) SDS-PAGE of Opti-prep purified supernatants (25 ng p24 equivalent) derived from 293T cells transfected with pQCXIP-FLAG-IFITM3 constructs, HIV-1 pNL4-3, and Vpr-Blam plasmid followed by immunoblotting with anti-FLAG M2 and anti-p24 Gag. Scale bar, 5 μm. (K) Opti-prep purified supernatants from (J) were incubated with SupT1 target cells for 4 hours and virus-cell fusion was scored by flow cytometry. The mean + SD of 3 experiments is shown.
Figure 1Average mature piRNA abundance relative to EV in EV and ints-11 RNAi samples normalized using miRNA-derived size factors. Error bars represent the standard error of the mean (5 replicates). Distributions of log2 fold change in piRNA abundance in ints-11 RNAi-treated nematodes compared to empty vector-treated nematodes in 5 replicates. Boxplots show the interquartile range with a line at the median; the whiskers extend to the greatest point no more than 1.5 times the interquartile range. Average piRNA precursor abundance relative to EV in EV and ints-11 RNAi samples, normalized to the total number of short capped RNA reads mapping to annotated WormBase TSSs (Chen et al., 2013). Error bars represent the standard error of the mean (5 replicates). Distributions of log2 fold change in piRNA precursor abundance in ints-11 RNAi-treated nematodes compared to empty vector-treated nematodes in 5 replicates. Boxplots show the interquartile range with a line at the median; the whiskers extend to the greatest point no more than 1.5 times the interquartile range. Distributions of piRNA precursor length in ints-11 RNAi-treated nematodes compared to empty vector-treated nematodes, obtained from deeply sequencing one pair of libraries (inserts sequenced up to 75nt). Distributions of total sequences (E) and total read counts (F) are shown.
Figure 8Model for the role of Gata6 in hair follicle progenitor cell survival through genome maintenance.
Figure 7(A) Representative confocal images of muscle-6/7 NMJ synapses of abdominal hemisegment A2 for the indicated genotypes labelled HRP (red) and with α-futsch (green) to reveal the neuronal membrane and Futsch protein level. Scale bar: 20 μm. (B) Quantification of normalized Futsch protein level at NMJ 6/7 in A3 of the indicated genotypes (n = 20). Bars show mean ± s.e.m. P values were determined with a Student's t-test. (p<0.05 = *; p<0.01 = **; p<0.0001=****). (C) Western Blot analysis of Futsch protein level in isolated third instar larval brains of different mutants compared to control. Numbers indicate normalized Futsch protein level in comparison to Tubulin levels. Bars are labelled with the number of replicates. (D) Quantification of futsch mRNA levels obtained from isolated 3rd instar larvae brains of the indicated genotypes via real-time PCR analysis. Bars show average ± SD of biological triplicates. Bars are labelled with the number of replicates. (E) Translating Ribosome affinity purification assay. Quantification of futsch mRNA levels upon immunoprecipitation of GFP-tagged Rpl10Ab or GFP in wildtype and Ythdf#0 3rd instar larvae with Real-time qPCR. Bars show average ± SD of technical triplicates. Bars are labelled with the number of replicates. (F) Quantification of normalized bouton number (total number of boutons/muscle surface area (μm2 × 1,000)) of NMJ 6/7 in hemisegment A3 of the indicated genotypes. Bars show mean ± s.e.m. Multiple comparisons were performed using one-way ANOVA with a post-hoc Sidak-Bonferroni correction (n.s. = not significant; p<0.01 = **; p<0.0001=****). Bars are labelled with the number of replicates.
Characterization of NF54, NF135 and NF175 A: Molecular characterization by PCR using established markers: MSP2 IC1, GLURP, MSP1 K1, MSP2 FC27, MSP1 R033, and MSP1 MAD20.Figure 2A: Molecular characterization by PCR using established markers: MSP2 IC1, GLURP, MSP1 K1, MSP2 FC27, MSP1 R033, and MSP1 MAD20. B: Markers as in A as well as the sizes for the three parasite strains. C: Expression of typical liver stage protein markers: Circumsporozoite protein (CSP) top left; Exported Protein 2 (EXP2) top right; Glyceraldehyde 3-phosphate dehydrogenase (GAPDH) bottom left; and Merozoite surface protein 1 (MSP1) bottom right. Scale bar is 25 µm.
PI(5)P Triggers Autophagy in the Absence of VPS34 Activation (A and B) Western blot analysis of LC3-II and tubulin levels and quantification of LC3-II/tubulin ratio in HeLa cells pretreated with Wm (200 nM, 2 hr), then loaded with 10 μM PI(5)P for 1 hr (in the presence of Wm and BAF) (mean ± SEM).Figure 5PI(5) P Triggers Autophagy in the Absence of VPS34 Activation(A and B) Western blot analysis of LC3-II and tubulin levels and quantification of LC3-II/tubulin ratio in HeLa cells pretreated with Wm (200 nM, 2 hr), then loaded with 10 μM PI(5) P for 1 hr (in the presence of Wm and BAF) (mean ± SEM). (C) Quantification of numbers of GFP-LC3 vesicles in HeLa cells stably expressing GFP-LC3 treated as in (A) and then shifted to starvation media (HBSS) or complete media (Basal) for 2 hr (in the presence of Wm) (mean ± SEM). (D) HeLa cells transfected with HA-ATG12 and ATG5 treated with 200 nM Wm as in (A) and subjected to western blot analysis with anti-HA antibody to detect free ATG12 and the ATG5-ATG12 complex (mean ± SEM). ATG5-ATG12 conjugation was checked for endogenous proteins in the same conditions using an anti-ATG12 antibody (bottom). (E and F) Western blot analysis of LC3-II and tubulin levels and quantification of LC3-II/tubulin ratio in HeLa cells transfected with control, PI5P4K2A, 2B, and 2C siRNA and treated with Wm (200 nM, 2 hr in the presence of BAF) (mean ± SEM). (G and H) Western blot analysis of free ATG12 and ATG5-ATG12 complex levels with anti-HA antibody in HeLa cells treated with control, PI5P4K2A, 2B, and 2C siRNA, transfected with HA-ATG12 and ATG5 for last 16 hr and treated with Wm for 2 hr (mean ± SEM). (I) HeLa cells stably expressing GFP-LC3 treated with control, PI5P4K2A, 2B, and 2C siRNA, were pretreated with Wm for 2 hr in complete medium and then shifted in HBSS media for 2 hr (in the presence of Wm) (mean ± SEM). (J and K) HeLa cells stably expressing GFP-LC3 were transfected for 30 hr with myc-tagged empty vector, myc-MTMR3WT, and myc-MTMR3C413S, incubated for 4 hr in HBSS in the presence of BAF and in the presence or absence of Wm. Cells were fixed, stained with anti-myc antibodies, and imaged by confocal microscope. Asterisks indicate transfected cells. Quantification of numbers of GFP-LC3vesicles per cell is shown in (K) (mean ± SEM). (J and K) HeLa cells stably expressing GFP-LC3 were transfected for 30 hr with myc-tagged empty vector, myc-MTMR3WT, and myc-MTMR3C413S, incubated for 4 hr in HBSS in the presence of BAF and in the presence or absence of Wm. Cells were fixed, stained with anti-myc antibodies, and imaged by confocal microscope. Asterisks indicate transfected cells. Quantification of numbers of GFP-LC3 vesicles per cell is shown in (K) (mean ± SEM). (L and M) HeLa cells transfected with GFP-PI5P4K2A, 2B, 2C, and RFP-LC3 for 30 hr were loaded with 10 μM PI(3) P for 1 hr in complete medium, and then fixed and imaged on confocal microscope. Bar, 10 μm. Quantification of cells (percentage of total) showing more than 10 RFP-LC3 vesicles in the different conditions from (J) is shown in (M); n = 200 cells (mean ± SEM). See also Figure S5.
Figure 6A Immunoblot of cell lysates from WT HAP1 cells and PTAR1 KO HAP1 cells using anti-Ykt6 antibody. Note the slight difference in the gel mobility of Ykt6 (arrows). B Prenylation status of rat brain Ykt6. Recombinant unprenyl Ykt6, Cys195-farnesyl Ykt6, Cys194/195-diprenyl Ykt6 samples (1 ng each), and rat brain cytosol were electrophoresed on a polyacrylamide gel using deoxycholate (DOC)-containing buffer and analyzed by immunoblotting with anti-Ykt6 antibody (DOC-PAGE; upper panel). The same samples were analyzed by conventional SDS-PAGE and immunoblotting with anti-Ykt6 antibody (SDS-PAGE; lower panel). C Prenylation status of Ykt6 in WT and PTAR1 KO cells. Recombinant Ykt6 samples and cell lysates of WT cells, PTAR1 KO cells, and PTAR1 KO cells stably expressing PTAR1 (KO + PTAR1) were separated by DOC-PAGE (upper) or SDS-PAGE (lower), and analyzed by immunoblotting with anti-Ykt6 antibody. D In vitro reconstitution of Ykt6 double prenylation. Dialyzed cytosol prepared from PTAR1 KO HAP1 cells was incubated at 37ºC for the indicated times with recombinant GGTase-III (100 nM) or RabGGTase (100 nM) in the absence or presence of GGPP (10 μM). After incubation, reaction products were separated by DOC-PAGE (upper) or SDS-PAGE (lower), and analyzed by immunoblotting with anti-Ykt6 antibody.
Figure 6Figure 6. RuBisCO activity and RuBisCO activation state is increased in the hda14 mutant under low light conditions. (A) RuBisCO initial and total activity in WT and hda14 in low light treated plants. Initial activity was measured directly upon extraction. For the total activity samples were incubated with H2CO3 for 3 minutes to fully carbamylate the active site of RuBisCO (n = 10, *t-test p-value < 0.05). (B) RuBisCO activation state. (C) ATPase activity of recombinant 6x-HisRCAß1 WT, K438Q and K438R with ATP and ADP/ATP= 0.11, respectively (n = 3, *t-test p-value < 0.05, +t-test p-value< 0.1).
Figure 1A. Schematic illustration of the syngeneic tumor model in which MC38 colon cancer cells were injected subcutaneously. 5-FU (25 mg/kg per dose) was administered intraperitoneally from day 6 post cancer cell injection, with one dose every two days. Tumor and spleen were harvested at 2 weeks. B. Tumors formed with WT MC38 cells were treated with PBS or 5-FU. Tumor volumes were quantified at the indicated days. N=5. C. STING-KO or sgRNA control (Ctrl) MC38 cells were treated in vitro with the indicated concentrations of 5-FU. Cell viability were determined using the CellTiter-Glo assay after two days, with normalized luminescence levels shown. N=3. Mice were injected with Ctrl or STING-KO MC38 cells, and treated with PBS or 5-FU. (D) Pictures of tumors and spleens from a representative experiment. Image panels were cropped from the same picture.  Mice were injected with Ctrl or STING-KO MC38 cells, and treated with PBS or 5-FU. (E) Tumor volumes were quantified at the indicated days post cancer cell injection. N=4 to N=5 as shown in (D). Mice were injected with Ctrl or STING-KO MC38 cells, and treated with PBS or 5-FU. (F) Tumor and spleen weights at the endpoint for (E), with each dot representing a mouse. WT MC38 cells were injected into STING+/+ or STING-/- mice, and treated with 5-FU or PBS following the schematics in (A). (G) Pictures of tumors and spleens from a representative experiment. Image panels were cropped from the same picture.  WT MC38 cells were injected into STING+/+ or STING-/- mice, and treated with 5-FU or PBS following the schematics in (A). (H) Tumor volumes were quantified at the indicated days post cancer cell injection. N=5. WT MC38 cells were injected into STING+/+ or STING-/- mice, and treated with 5-FU or PBS following the schematics in (A). (I) Tumor and spleen weights at the endpoint for (H), with each dot representing a mouse.
Figure 3A, B. Representative images showing endogenous hTrmt13 (A) and HA-tagged hTrmt13 (HA-hTrmt13, B) subcellular localization by confocal microscopy analysis. DAPI staining was included to visualize the cell nucleus (Blue). C. Representative confocal microscopy images showing the subcellular localization of hTrmt13 in MDA-MB-231 cells cultured with 10% FBS (left) or no FBS (right). DAPI staining was included to visualize the cell nucleus (Blue). D. The distribution of hTrmt13 analyzed by subcellular fraction assay. β-actin as a marker of cytosol, Lamin A/C as a marker of nucleoplasm, Histone H3 as a marker of chromatin. E. Confocal microscopy images of MDA-MB-231 cells were co-immunolabeled with antibodies against hTrmt13 and histone H3 (left). The images were captured by GE DeltaVision OMX SR. Co-localization of H3 and hTrmt13 was analyzed by ImageJ (right, n = 20 cells, center line, the median of the data). DAPI staining was included to visualize the cell nucleus (Blue). F. Schematic diagram illustrating the principle of nucleic acids-IP performed in nuclear extract and cytoplasm without adding RNase or DNase. G. hTrmt13 bound nucleic acids from the cytosol and nuclear extract were stained by GoldView. Transcribed HctRNAGly(GCC) was used as a size marker for tRNAs. H. Genomic DNA was sheared to fragments by Biorupt (left). EMSA analysis of hTrmt13 and DNA fragments (15 ng/μL DNA was used). I. The binding profiles of hTrmt13, H3K4me3, H3K27Ac, and other transcription factors (MYC, E2F1, TP53, FOSL1, EZH2, FOXM1) from ChIP-seq and ATAC-seq on representative target genes TGFB1, MEF2A, and PKN2. hTrmt13 track were average across 2 biological replicates. J. Genomic distribution of hTrmt13 reproducible (N=2) binding sites determined by ChIP-seq analysis in MDA-MB-231 cells. k, hTrmt13 binding de novo motif for reproducible (N=2) binding sites. L. hTrmt13 reproducible (N=2) binding sites enriched for known motifs for FOSL1, FOSL2, JunB, BATF, and AP-1. M. The binding affinities of wild type and mutant hTrmt13s for synthesized DNA (0.25 μM) analyzed by EMSA. The concentration of hTrmt13-dZ2 or hTrmt13 used in the system was 1 μM. N. EMSA of hTrmt13 for synthesized DNA (0.25 μM) carrying consensus motif or mutated motif.
TEAD and AP1 activation are distinctively controlled by nuclear shape.Figure 4Quantifications of YAP activity in Ctrl, Blebb and Blebb + AP conditions, treated or not with importazol (nucleo-cytoplasmic ratio). Immunoblots of p-YAP (ser127), YAP, p-LATS (Thr1079), LATS, p-MST (Thr1083), MST, Kibra, PTPN14 and GADPH for Ctrl, Blebb and Blebb + AP conditions. Corresponding quantification of p-YAP (Ser125) relative to Ctrl and normalized to GAPDH. Corresponding quantification of p-LATS (thr1079) relative to Ctrl and normalized to GAPDH. Quantifications of p-Jun Ser63 nuclear intensity in Ctrl, Blebb, and Blebb + AP conditions for cells treated with DMSO, SB203580, a MAP Kinase inhibitor and SP600125, a JNK inhibitor. Representative cells stained for p-Jun Ser63 (magenta) and for DNA (cyan) in Ctrl, Blebb and Blebb + AP conditions for cells depleted or not for lamin-A/C. Scale bar = 10 µm. Corresponding quantification of p-Jun Ser63 nuclear intensity.