EMBO/sd-character-level-ner huggingface dataset:
G, H. Average read profiles on two groups of RefSeq genes, in the size-range 4 to 15kb, containing the highest (G) or lowest (H) CACACA motif density.
Immunoblot of lysates prepared from the indicated SAMHD1-proficient (+/+), deficient (-/-) and rescue (WT, H233A) cell line pairs with the indicated antibodies. Representative of 2 independent experiments.
(d) Quantification of adipose tissue volume in larvae carrying tumors of RasV12; ctrl at day 6 and RasV12; scrib-/-at days 6 and day 10.
F: Rates of PB1 emission and normal spindle formation in oocytes ovulated by WT and Cnot6l-/- female mice. The numbers of analyzed oocytes were indicated (n)
Confocal images of control and MCOLN1 shRNA-expressing HN31 cells treated with the cholesterol sensor, filipin. Scale bar (10μm) shown in the panel on the top applies to both panels. Bar graph showing the quantification of the internalized filipin staining in HN31 cells after MCOLN1 knockdown. All values represent mean±SEM. Data points represent values from biological replicates. Statistical test employed was Student's t-test.
F. The decrease in MCH mRNA levels observed 18h after icv LPS injection is partly abolished by the CCR2 antagonist INCB3344 (10μM) and in CCR2 KO mice, as shown by real-time PCR analysis for MCH in the hypothalamus. Results were normalized to values in icv saline injected WT mice (n=6 per group). Data are expressed as means ± SEM. ** or $$ p<0.01. * saline vs icv LPS injection in WT mice. $ icv LPS injection in WT mice vs LPS+INCB3344 icv injection in WT mice or icv LPS injection in CCR2 KO mice.
B, Impact of the 5' DSB/repeat distance on RMD frequency when the 3' DSB/repeat distance is fixed. RMD assays were performed using mES cells carrying the RMD-GFP reporter, and RMD frequency was determined by the number of GFP+ cells normalized to transfection efficiency. The distance between the sgRNA targeting sites and the repeats is indicated.
(A) Sectioned third instar OreR larva stained with Periodic acid-Schiff (PAS). The muscles, but not the fat body, are stained purple, indicating high levels of glycogen (m, muscle; bw, body wall; fb, fat body).
E Detection of the time-dependent accumulation of AMPylated BiPT229A-V461F in radioactive reactions, containing [α-32P]-ATP and the indicated FICD proteins, in the presence of excess trap. At the specified time-points samples were taken and analysed by SDS-PAGE. The autoradiograph (32P) illustrates the radioactive signals, which represent AMPylated BiP. Proteins were visualised by Coomassie staining. The radioactive signals were quantified and presented in the graph below. Mean values ± SD of three independent experiments are shown.
C-D. Volcano plot showing differential expression (DE) between Alkal2 and MYCN tumours. Differentially expressed genes are shown in blue (DE in Alkal2 tumours only) or black (DE in both Alkal2 and AlkF1174S tumours, as shown in (D)). Top ranked genes labelled. Dashed lines represent DE cut-offs.
A, B Immunohistological staining of liver sections from P240 PR-SET7ΔHepA mice and control littermates (WT) with Afp and HNF4 (A) or Ki-67 and albumin (Alb) (B) antibodies.
(f) Pearson correlation coefficient of mean nuclear Venus::HES5 intensity in relationship to distance; red dots indicate average Venus::HES5 correlation per slice of 12 slices from 3 experiments with corresponding red line indicating polynomial fit (order 2); gray dots with black line indicate correlations and polynomial fit from 5 randomizations of intensities analysed in the same way (see Materials and Methods).
C,D Summary graphs of the membrane distribution of Syx1 (C) and Munc18-1 WT or mutants (D) quantified by the ratio of fluorescence at the plasma membrane (PM) to the total fluorescence in the whole cell (total). DiD is the PM marker. Numbers of pictures analyzed are shown as follows: no Syx1, n = 14; no M18, n = 15; WT, n = 24; QKAA, n = 23; KKEE, n = 12; L348R, n = 22.
C In situ hybridisation of ptch2, nkx2.2a and olig2 transcripts in wild type, MZgrk2 and grk2-GFP injected MZgrk2 24hpf embryos. Each panel shows a full view of the embryo on the left and a cross sectional view of a somite on the right. (n=30 for each sample). Scale bars, 200μm (whole mounts), 50μm (sections).
H Size-exclusion profiles for apo and iron-loaded forms of DJA6. 1 mM EDTA, purified DJA6 after incubated with EDTA at a final concentration of 1 mM; 20 μM or 100 μM Ferrous, apo-DJA6 after incubated with Fe(NH4)2(SO4)2 at a final concentration of 20 μM or 100 μM, respectively. Positions of theoretical tetramer and dimer are indicated by dotted lines. Images of the Coomassie Brilliant Blue-stained gel after SEC elution are shown.
(B) sRNA-seq reads profile of EVD expressed from 35S:EVDWT in WT (black) or rdr6 (red). (RPM) reads per million. Positions are indicated in nucleotides (nt) from the start of the 35S sequence. Dashed vertical lines: shGAG and GAG-POL 3' ends.
NlpI has different binding sites for MepS and PBP7 as shown by the ability of labelled MepS to bind preformed NlpI-PBP7 (D) complexes by a fixed concentration MST assay. Values are mean ± standard deviation of 3-6 independent experiments. To calculate significance, the data were fit using a linear model. Calculated means were compared using Tukey's HSD test, resulting in p-values corrected for multiple testing. Relevant p-values are highlighted directly in the figure (*<0.05; **<0.01, ***<0.001)
(D) Average of ER-mitochondria distance (n= 111-168 mitochondria). *p<0.05, Mann-Whitney test. Data are presented as mean ± SEM. (E) Quantification of the percentage of mitochondria in contact with the ER distributed at the indicated distance range (n= 3 independent experiments in which were analyzed 627-779 mitochondria from 68-98 cells). Data are presented as mean ± SEM. *p<0.05, two-tailed Student's t-test. Data information: Note that experiments were performed at the same time, so the values of WT and Mfn2 KO are the same but the figures have been split in two for the sake of linearity.
E LC-MS/MS analysis of d3-m6A/A levels in 18S rRNA oligos after being incubated with recombinant WT or catalytically mutant mouse METTL5 proteins in vitro. No enzyme group was used as a negative control.
A. Confocal microscopy of tau (red pseudocolor) and Hsp70 (green pseudocolor) colocalization (merged image) levels in wild type primary neurons overexpressing GFP-AAV9 or FLAG-DnaJC5-AAV9. Boxed area are 5X digital zoom on a 60X lens shown below. Scale bars are 20 µm and 5 µm as indicated. Only neurons overexpressing virus were imaged.B. Quantification of tau-Hsp70 co-localization (mean Pearson's coefficient ± SEM, n=18, *p <0.05, one-way analysis of variance with Tukey's multiple comparison post-hoc analysis).
(j) CTR and RAB2A MCF10A cells were transfected with Str-KDEL_SBP-EGFP-E-cadherin (green). At the indicated time points after the addition of biotin, cells were fixed, and stained without permeabilization with anti-GFP antibody, followed by Alexa568-conjugated secondary antibody (red). The cell surface and total levels of SBP-EGFP-E-cadherin at 60 min after the addition of biotin were quantified using ImageJ software on non-saturated images, and surface/total ratio was expressed as a surface GFP index (relative to control cells). Bar, 20 µm. ***, P < 0.001.
C. Western blot analysis of Tace in Mtmr2-/- sciatic nerves at P5, P10, and P20. Representative of two independent experiments.
A. Violin plot showing the expression of MCT family members (monocarboxylate transporters) in brain pericytes from scRNAseq database (Brain cell atlas; 1,100 endothelial cells)
CD3+ T cells (upper) or CD4+ and CD8+ T cells (lower) in the spleens of skin-graft recipient mice were analyzed by FACS.
(F-G) K413 in IDH2 may be a key succinylation site that is directly targeted by SIRT5. The indicated Flag-tagged IDH2 proteins were overexpressed in HEK293T cells with stable SIRT5 knockdown or SIRT5 co-overexpression. Wild-type and mutant IDH2 proteins were purified by Flag beads and eluted with Flag peptide, followed by measurement of IDH2 enzyme activity. The results are average ± SD of 3 independent experiments *p<0.05, n.s.: not significant (two-tailed unpaired t-test).
A Topology of functional, binding and regulatory motifs and domains in the proposed endocytic coat architecture. Known motifs and domains of indicated coat proteins were classified according to their main function and color-coded as shown on the right. Ark1/Prk1 phosphorylation sites were taken from (Huang et al., 2003).
B Wild‐type (WT) and USP45 knockout (KO) U2OS cells were lysed and subjected to immunoblotting as in (A).
C, D) Coronal sections from wt and Sfrp1-/- mouse brains three days after infusion of saline or LPS, immunostained for GFAP (cyan, C) or CD45 (magenta, D). The images are high power views of the somatosensory cortex (lower power view in Fig EV1D). Scale bar 60μm. E, F) The graphs show the normalised levels of immunoreactivity (IR) for GFAP (E) and CD45 (F, P=0,006) present in cortical sections (n=24 acquisitions white dots; from N=3 animals, black dots, per group) from wt and Sfrp1-/- animals infused with saline or LPS. Bars represent Standard Error. Statistical significance calculated per biological replicas is indicated in green and that based on number of acquisitions in black. ** or ## P<0.01; *** or ### P<0.001; **** or #### P<0.0001 by two-way ANOVA followed by Bonferroni's multiple comparisons test. * and # indicate significance between genotypes and treatments, respectively.
M. Quantification of the number of esg-GFP+ cells, Dl+ cells, and pH3+ cells in experiments I-L. n is indicated. The numbers of quantified guts from left to right are 15, 17, 18, 17, 15, 17, 18, 17, 15, 17, 18, and 17. Data information: Error bars represent SDs. Student's t-tests, *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001, and non-significant (NS) represents p > 0.05.
A Electron microscopy analyses to see autophagic bodies in the vacuoles under both growth and nitrogen starvation conditions. Autophagic bodies typically contain cytosolic ribosomes. pep4∆prb1∆ and pep4∆prb1∆atg2∆ cells were grown in SD to mid‐log phase (OD600 = 1) and transferred to SD(‐N) for 5 h. The cells were examined by transmission electron microscopy as described in . Scale bar, 500 nm.
(F) Kaplan-Meier plot analysis of the disease-specific survival of 85 TNBC breast cancer patients with low or high expression of MIG-6.
C. GSDMD at a concentration of 1 μM was incubated at room temperature with 5 nM caspase-1 and liposomes composed of 4 mM DMPC or polar lipid extract derived from E. coli. After 2 h, the lipid fraction (L) was separated from the supernatant (S) by ultracentrifugation at 4C for 1 h at 120,000g.
(d) MCF-7 side-population (SP) cells have a significantly higher percentage of MBd+ cells over the non-side-population (MP; P=0.0015, n=3; data are mean±s.d.).
HPV16E6 protein prevented YAP protein from degradation. Confluent HT3 cells were starved for 4 h before treatment with or without CHX (20 μg/ml), HPV16E6, or CHX combined with HPV16E6 for 8 h. Treatment of starved HT3 cells with HPV16E6 (400 nM) for 8 h suppressed degradation of YAP, but not EGFR protein.Quantitative data showing relative YAP protein levels in (D). Protein levels were normalized with β-actin and presented as ratios relative to that of control. Each bar represents mean ± SEM (n = 4). Bars with different letters are significantly different from each other (Ctrl vs. CHX, P = 0.0003; Ctrl vs. CHX+E6, P = 0.2665; Ctrl vs. E6, P = 0.0258).
1205Lu cells exposed to 3T3-F442A EV (3T3-EV) and treated, or not, with Mdivi-1, were stained with a MitoTracker probe, fixed and observed by confocal microscopy. Arrows indicate the presence of mitochondria in membrane protrusions. On the images to the right, the outline of cells is shown in dotted lines. Scale bars: 20µm. Right panel, quantification of the distance between mitochondria and nuclei (n=15).
F,G) Immunocytochemistry of DIV4 primary hippocampal neurons from wild type, Ttll1-/- and Ttll7‑/- mice. Microtubules in Ttll1-/- cells have reduced levels of polyE signal, indicative of lower α-tubulin polyglutamylation (F), Ttll7‑/- neurons are specifically reduced in β‑monoE reactivity, indicating the lack of β‑tubulin glutamylation (G).
WT and Mfn2 KO MEFs were co-transfected with plasmids encoding: H) LAR-GECO1 and ER-Mfn2 or control (CT) plasmid. ER Ca2+ levels were determined. ER Ca2+ release was induced with caffeine (20 mM) as indicated (n= 35 cells from 4 independent experiments). Data are presented as mean ± SEM Data information: *p<0.05, one-way ANOVA followed by Tukey's post hoc test. Note that experiments were performed at the same time, so the values of WT and Mfn2 KO transfected with control plasmid are the same but the figures have been split in two for the sake of linearity.
A. Schematic illustrating the preparation of samples for media analysis by dot blot/Western blot or mass spectroscopy.
B. Genetic interactions of nlpI with EPase genes. Strains were arrayed using a Rotor HDA replicator on Lennox LB agar plates and incubated for 12 h at 37°C. Each plate contained 384 colonies, 96 from the wildtype, single mutants and double mutants. An example of a 384 plate is shown. Double mutants were made twice, swapping the resistance markers to the two single mutants. Colony integral opacity was quantified as a fitness readout, using the image analysis software Iris (Kritikos et al., 2017). Bar plots shows the averaged values of 2 biological experiments, each having 96 technical replicates (i=2, n=192). The error bars represent the 95% confidence interval.
CSF extract was supplemented with Myc-XErp1 DSG- DSA- ZBR- (S33N S38N S284N S288N C583A) IVT and as indicated treated with DMSO or CsA. Both reactions were treated with calcium and samples were taken at the indicated time points. Samples were immunoblotted for the Myc-tag, cyclin B2 and α-tubulin. A pseudocoloured representation of the Myc immunoblot is shown in Fig EV2A. Arrow marks the meiotic phosphorylation state of Myc-XErp1. Asterisk indicates unspecific bands.
Real-time qPCR analysis of fetal organoids cultured for one month showing decrease in relative expression of indicated neonatal markers A Ass1, B Blimp-1, C FcRn and D Lct and increase of the adult markers E Sis, F Treh and G Arg2 (n=3 individual wells from single organoid culture (see material and methods); experiment was repeated in four to eight independent organoid cultures with similar results).
N=Native fold, U=unfolded state, A=aggregated state, T=transition state. A) Effects of GK placement on the energetics of the native, unfolded and aggregated states. GKs generally come with a cost to the native fold stability, with negative GKs being more destabilizing on average owing to their shorter sidechains and resulting increased burial costs (a). On the other hand, acidic residues more effectively destabilize the aggregated state than basic residues (b). Consequently, structural constraints in the native state may lead to APRs that are inadequately protected by their basic GKs. Hsp70 specifically recognizes these APRs through their inadequate GKs, and effectively compensates for the difference in destabilization of the aggregated state between basic and acidic residues (c).
(A) GFP-LC3 CHO cells were preincubated with 1 mM dybutiryl cAMP (+dbcAMP, panels d, e, and f) for 30 min and then were treated for 4 h with 10 µg/ml of α-hemolysin (Hla) or subjected to starvation conditions (Stv) in the continuous presence of dbcAMP. Cells without any treatment were used as control (−dbcAMP, panels a, b, and c). Cells were immediately analyzed by confocal microscopy. A quantification of the percentage of cells presenting LC3 puncta (i.e., stimulated cells) upon incubation with the different conditions is shown in the right panel (n = 100 cells/condition, * p<0.05 paired Student's t-test). These data are representative of three independent experiments.
B. Quantification of excretion numbers of Drosophila with indicated genotypes. Excretions are quantified in 30 fields for each group of 12 Drosophila. Tests were repeated as three independent experiments. Data information: Error bars represent SDs. p-values for lifespan curves (C) were calculated by the log-rank test. The statistical tests used in other panels were student's t-tests. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001, and non-significant (NS) represents p > 0.05.
Repsox (1 μM) was used on day 2-7 to inhibit early EMT during reprogramming with 5C medium. The expression of glycolysis markers was determined on day 6 with qPCR (G).
E. Food intake measured using CAFE assay of Drosophila carrying esgts-GAL4-driven UAS-lacZ (control), Las RNAi, or Las RNAi with ALA administration. Error bars show the SD of three independent experiments. Data information: The statistical tests used in other panels were student's t-tests. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001, and non-significant (NS) represents p > 0.05.
(C) WT (n = 15), Irgm2-/- (n = 12), and panIrgm-/- (n = 13) mice were injected i.p. with LPS (8 mg/kg). Serum was collected 4 hpi and concentration of IL-1β and TNFα was measured via ELISA. Data shown are from 3 pooled experiments.
B. cdc20ts-mAR PDMC1-AMA1 PHSL1-CDH1 cells were released from the metaphase I-arrest with CuSO4 (t = 0) at 25°C. Subsequent incubation at 25°C allows progression through meiosis II (left), while a shift to 37°C at t = 50 min causes arrest at metaphase II (right). Mpc70-GFP and the PSM marker RFP-Spo2051-91 were imaged in cells fixed at the indicated times. RFP-Spo2051-91 is expressed from an episomal plasmid (Diamond et al., 2009). Images represent ≥ 90% of cells harboring the plasmid. Scale bar, 2 μm. Data information: data are representative of two independent experiments.
C Detection of free RNA. Wild‐type, atg2∆, rny1∆, and rny1∆atg2∆ cells grown in SD (left) or SD(‐N) for 2 h (right) were stained with FM4‐64 and GR Green and observed by fluorescence microscopy. Scale bar, 5 μm.
G. Survival (percentage) of female W1118 Drosophila with and without supplementation of ALA as indicated. The numbers of quantified flies: 100 (W1118 + 0.5 mM ALA) and 100 (W1118 + 0 mM ALA). Three independent experiments were conducted. Data information: Error bars represent SDs. p-values for lifespan curves were calculated by the log-rank test. The statistical tests used in other panels were student's t-tests. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001, and non-significant (NS) represents p > 0.05.
I. Isotope carbon tracing with [13C3]-lactate treatment (20mM, 24hr) in siCON or siMCT12-treated HBVPs showing the extent of downstream metabolites and incorporation rate of labeled carbons. (F-6-P : Fructose 6-phosphate, G-3-P : Glycerol-3-phosphate, Lac : Lactate, Cit : Isocitrate and aKG : α-ketoglutarate) (n=3 from independent experiments. Error bars indicate the standard error of the mean (SEM) from unpaired Student's t test.
A) Microtubules perturbations impact the trancriptome of differentiating cells. Volcano-plot representation of down-regulated (blue) and up-regulated genes (red) in non-treated/control cells relative to ciliobrevin-D treated cells. Each circle represents the fold change (FC) of the quantity of mRNA and the significance of the change based on three independent replicates. Expressed genes with a false discovery rate (FDR) lower than 0.05 are shown in grey, whereas genes with a FDR greater than 0.05 are shown in blue or red. Genes significantly mis-regulated i.e. with an absolute value of FC higher than 1.8 are shown as full disks. Names of genes of interest are indicated within the graph. Genes that are in the signature of the myeloid-differentiation pathway are shown in magenta. Genes that are the signature of the megakaryocyte-differentiation pathway are shown in green. B) Heatmap representation and hierarchical clustering of gene expression changes between non-treated and ciliobrevin-D-treated cells collected from 3 donors in each condition. The color code corresponds to a log2 scale of the differential expression levels.
b) Histograms of GFP expression (x-axis) are presented from A549 cells transduced with the different viral vectors.
Immunostaining for Pax7 together with Laminin was performed on the TA muscles 3 days post-CTX injury. Scale bar = 100 µm. Quantifications of the numbers of Pax7+ SCs per area are shown on the right (n=3 mice, each).
Quantificaiton of dCK phosphorylation at serine-74 (S74) with respect to total dCK in SAMHD1+/+ THP-1 cells treated with either solvent, 500 nM ara-C or a combination of 500 nM ara-C and an RNRi (HU, 50 µM; dF-dC, 10 nM; 3-AP, 150 nM) for 24 h. Circles and squares, columns and error bars correspond to individual measurements, means and s.e.m. of one representative out of two independent experiments performed in triplicates.
ChIP-qPCR was performed to show the enrichment of H3K27 me3 (R) binding on selected mitochondrial genes.
A-C Wild‐type, atg2Δ, pho8Δ, and rny1Δ cells were grown in SD to mid‐log phase and transferred to SD(‐N) at time 0. At the indicated time points, 3′‐NMPs (C) were analyzed by LC/MS as described in . The results are presented as normalized intensities on the basis of the peak height of each metabolite in wild‐type cells. Note that 3′‐NMPs in pho8Δ are presented in a different scale. All data are means of triplicates. The error bars represent the standard deviation.
(e AFOG staining of heart sections from 60 dpci pkma2+/-; pkmb+/- and pkma2-/-; pkmb+/- animals; black dashed lines outline the scar area; scar area measured on the right (n = 3-5 ventricles of each genotype).
A, B Western blot analyses of global protein O-GlcNAc levels in left ventricles of mice with either (A) cardiac-specific LXRα overexpression or (B) LXRα deficiency and subjected to 5 weeks TAC.
(H) Genomic snapshots depict RNAseq profiles of the selected glycolytic genes (Hk2, Glut1, Ldha, Pdk1, Eno1) down-regulated in YY1iKO vs Ctrl ASCs.
F. Rosa26_Alkal2;Th-MYCN tumours from lorlatinib or vehicle controls were analysed for pHis. A representative field of view for each tumour at 40x (175740μm2) were manually counted. Data shown represents mean ± 95% CI. p=0.0286; Mann-Whitney test.
Immunoblot showing co-immunoprecipitation of Dpb11 with Sgs1. Both DPB11-3xHA and SGS11-647-5xFLAG were expressed at their endogenous loci.
(B) Percentages (mean ± S.E.M.) of tumor burden in mammary glands of untreated (control) and sunitinib-treated NeuT;Apln+/+ and NeuT;Apln-/- mice, assessed 4 weeks after tumor onset. NeuT;Apln+/+ Control (n=6), NeuT;Apln-/- Control (n=8), NeuT;Apln+/+ Sunitinib (n=5), NeuT;Apln-/- Sunitinib (n=5); *P<0.05; ***P<0.001; one-way ANOVA. Representative H&E images are shown for each genotype in the right panels. Scale bars = 1000 μm (large panels) and 50 μm (insets).
D. Average cell size of esg-expressing cells (μm²); note the increased cell size of Dronc fully mutant progenitor cells (-/-) at 3d (**; P = 0.0014) and 7d (***, P = 0.0007) post temperature shift at 29 Co (Quantifications were made using N ≥ 2 biological replicates; unpaired two-tailed parametric t-test, 3d +/- n = 19, 3d -/- n = 28, 7d +/- n = 29, 7d -/- n = 28).
Primary ILC3 cells were sorted from LPs and were stimulated with IL-7 in combined with FIZC and AhR Inh. Cells without stimulation were used as control (None). Production of IL-22 was detected by ELISA.
D. Quantification of the pericytes coverage using CD13 immunostaining in the brain vasculature of WT (blue) and Glut1iECKO (orange) mice. (n=7 from 3 independent experiments. Error bars indicate the standard error of the mean (SEM) from unpaired Student's t test.) E. Quantification of the smooth muscle cell coverage using αSMA immunostaining in the brain vasculature of WT (blue) and Glut1iECKO (orange) mice. (n=13 from 3 independent experiments. Error bars indicate the standard error of the mean (SEM) from unpaired Student's t test.)
Apoptosis was determined by quantification of TUNEL-positive cells per field of view (FOV) in MDA231-LM2 tumors (from the animals described in (C)) treated with JNKi and/or PAX. Values are means from 6 mice per condition ± SEM.