EMBO/sd-character-level-ner huggingface dataset:
H Coimmunoprecipitation analysis of FADD and RIP140 in 293T cells. As indicated, lysates from the cells transfected with plasmids encoding Flag-RIP140, HA-FADD-A or HA-FADD-D were subjected to immunoprecipitation with an anti-Flag antibody. FADD-A and FADD-D were detected using an anti-HA antibody. Data shown are representative of three independent experiments having similar results.
b, Immunoblot of −KRas cells treated or not with oligomycin (Oli) probed with anti-Thr-172-phospho-AMPK and &amp;amp;bgr;-actin antibodies for three independent tumours.
E-G) Representative Western blot analysis of the protein level (E) and quantification of the translation efficiency of S2R+ cells transfected with the wildtype or mutated futsch reporter constructs upon control or Mettl3 KD (F) or Ythdf KD (G). LacZ dsRNA was used for the control KD. The translation efficiency was calculated as the ratio of the relative GFP protein level and GFP mRNA level. mCherry served as a normalization control for both the protein and mRNA level. Bars show mean ± s.e.m of biological triplicates. P values were determined with a Student's t-test. (n.s. = not significant; p<0.05 = *; p<0.01 = **).
L. NESTIN and GFP immunostaining of Ctrl, NDIME-/-, and NDIME-/-+Rs26-NDIME cells at day 5 of neural differentiation. Scale bars, 100 µm.
1 or 2 μM of GST or GST-TRF1 were incubated with the indicated concentrations of human mTOR kinase
(E) Graph shows the percentage of Pax6+ cells among GFP+ cells (n = 5 each group; two-tailed
D and E. HeLa cells were transfected with the indicated FUNDC1-MYC constructs for 24 h. Cell lysates were immunoprecipitated using anti-Calnexin and immunoblotted using anti-MYC and anti-Calnexin antibodies.
A-C: Volcano plot showing gene expression trends at 8h MMA treatment. Dashed lines represent a 2 log2FC (drug over control) (A) or 1.5 log2FC (drug over control) (B) and 10e-7 q-value threshold. Quantification of induced and repressed genes in (C). Labels indicate names of genes that meet the logFC and q-value threshold.
(F) Detection of ssDNA breaks by DNA break end labeling. The DNA breaks were end-labeled in situ by incorporating biotin-16-dUTP and TdT. The DNA was fragmented and labeled fragments were enriched by streptavidin pull down, followed by quantification by qPCR. The signal from the β₂-Macroglobulin (β₂m) locus served as a negative control. The values are presented as mean ± sd (n=3). Statistical significance was assessed by two tailed unpaired Student's t-test (*** p ≤ 0.001). The two DNA break assays were illustrated in Supplementary (Appendix Fig S3E, B and F).
A. Genome browser view of AC and DMZ accessibility profiles for ectoderm-expressed (tfap2a and grhl3) and organizer-expressed (gsc and chrd) marker genes.
A. WT or LAT-deficient Jurkat cells (Jcam2.5) expressing CAR-GFP were stimulated on SLB-coated with OKT3 (anti-TCR antibody) or CD19 (CAR antigen); in both conditions, the lipid bilayers were also coated with ICAM-1 to facilitate cell attachment. TIRF microscopy revealed clustering of Ax647-labeled streptavidin-Biotin-OKT3 or CD19, which serves as a probe for TCR or CAR respectively. Scale bar, 2 µm. B. Quantification of clustering as normalized variance. n = 100 cells. Shown are the mean ± SD. Statistical test: unpaired two-tailed t-test. **** p < 0.0001. ** 0.005 < p < 0.05.
(e) As in d, using lysates from U2OS cells expressing Flag-Parkin and T7-Fbxo7, either full length (1-522) or a C-terminal deletion of the proline-rich region (1-398). * indicates IgG heavy chain.
F) Quantification graph of the velocity of EB1-GFP comet movement in the protrusion of qNSCs at 0 h ALH, 6 h ALH and 10 h ALH, as well as in interphase NSCs (aNSCs) at 72 h ALH. n=250 for 0 h qNSCs (anterograde); n=30 for 0 h qNSCs (retrograde); n=211 for 6 h qNSCs (anterograde); n=24 for 6 h qNSCs (retrograde); n=171 for 10 h qNSCs (anterograde); n=103 for interphase NSCs (anterograde). ****p<0.0001; * p=0.0285
A shERWOOD-designed shRNAmir targeting mouse transcriptional factor Tbx21 and a control shRNAmir targeting mouse CD4 were cloned into both miR-AB and miR-E retroviral vectors with Venus or Ametrine as reporter. The packaged viruses were used for transduction of primary mouse P14 TCR transgenic CD8+ T cells, which were used for evaluation of CD8+ T cells differentiation in vivo as described previously(Wang
(I) mRNA expression analyses of TECR and TECRL by RT-qPCR in human tissues demonstrates preferential expression of Tecrl in the heart; [Bone marrow (Bone M.), heart, skeletal muscle (Sk.Muscle), uterus, liver, spleen, thymus, thyroid, prostate, brain, lung, small intestine (Small I.) and colon]
B Equal amounts of total RNA isolated from spermatocytes of Miwi+/-;Dicerf/+;Stra8-Cre, Miwi-/-;Dicerf/+;Stra8-Cre, Miwi+/-;Dicerf/-;Stra8-Cre and Miwi-/-;Dicerf/-;Stra8-Cre mice, were resolved on a 1% agarose gel. The gel was subjected to Northern blot analysis using probes specific for the sense and antisense sequences of major and minor satellites. The blot was re-hybridized with a 28S rRNA-complementary probe to demonstrate equal loading (bottom on the left).
A-B'') Representative images showing that Lsd2YFP is localised to the surface of LDs (yellow arrows).
C: Schematic highlighting whether a mitotic interaction was previously known between FAM83D and the interacting proteins
HO-1-/- mice have more ECs and less CARs (p = 0.053) in BM, while frequency of PaS does not differ.
(g-i) Fluorescent microscopy of endogenous LINE-1 RNA in cells transfected with GFP-DCP1A (g), GFP-GE-1 (h) or GFP-TIA-1 (i). All experiments were performed with a minimum of three biological replicates.
Snapshots of Movie EV5. Time Lapse of embryonic organotypic slice from a CD41:YFP: H2B-GFP reporter mouse showing the recruitment of CD41:YFP negative cells (magenta central point-tracking analyses) migrating towards the IAHC (CD41:YFP+ cells). Green central point and relative tracking identify cells with non-directional movement. Scale bar: 5μm. Time expressed in hh:mm:ss.
(D) BMDMs were pretreated with rRv2626c-WT, rRv2626c-CA, or rRv2626c-DN for 1 h, and stimulated with 100 ng/ml LPS for 18 h. Culture supernatants were harvested, and the levels of TNF-α, IL-6, IL-12p40, and IL-10 were measured by ELISA.
3′ UTR of the Tnfrsf1a gene of B mice. The single nucleotide polymorphisms (SNPs) found in the sequence of S mice are shown in red. miR-511 was predicted by the program MicroSNiPer (http://cbdb.nimh.nih.gov/microsniper/) to have two target sequences, shown in bold and underlined. The most 3′ target sequence contains two nucleotide differences (in red) in S (lower blue panels) compared to B (upper blue panels), leading to different base pairing with miR-511, as shown in the lower panel.
FANCD2 ubiquitination cannot robustly enhance ID2-DNA binding when complexed with IR1285Q, as determined by EMSAs. IRDye700-labelled dsDNA (at 2 nM) was incubated with indicated amounts of ID2 (His6-FANCI or His6-FANCIR1285Q mixed with non-ubiquitinated or ubiquitinated FLAG-FANCD2. Mixes were ran on non-denaturing gels, and the resolved free- and protein-bound DNA bands were visualized using an infrared scanner. Gels shown are representative of two replicate experiments.
(U) Model for the role of autophagy in modulating the miRNA pathway. co‐IP, coimmunoprecipitation; GFP, green fluorescent protein; MBP, maltose‐binding protein; miRNA, microRNA; IP, immunoprecipitant; RISC, RNA‐induced silencing complex; s.d., standard deviation.
(F) Percentage of cells infected with WT Listeria, in the presence (black bars) or absence (white bars) of rapamycin, displaying one or several GFP‐LC3+ Listeria autophagosomes. Values are means±s.e.m. n=2. Scale bars: 5 μm.Source data for this figure is available on the online supplementary information page.
Plasmid contents of the strains carrying the sRNA libraries before selection (input) and after consecutive ethanol treatments (Sel1, Sel2, Sel3) were analyzed using high-throughput sequencing. Relative library complexity (left y-axis) was determined by counting sequence variants present in the normalized samples. To test for the enrichment of possible sequence motifs, the sequence variants present in each sample were counted and normalized for sequencing depth. The resulting data were analyzed for the enrichment of the conserved CRCUGCUUUU motif (right y-axis).
B, C Relative abundance of sgRNAs in cell populations with high versus low signal were visualized as log2 fold chances for basal as well as activated autophagy conditions for (B) p62 or (C) NDP52. Entire data is reported in Dataset EV1
(c) Altering the shape of PE to reduce local membrane defects inhibits the lipidation reaction. In vitro coupling reactions of GL1 were run on liposomes of three different sizes (400 nm and 50 nm extruded liposomes and sonicated liposomes) and of three different lipid compositions. Lipid compositions include 30 mol% DPPE, DSPE or DOPE (see Methods for full definitions), 10% bl-PI, and POPC. Total lipid = 2 mM. Quantification is of n = 3 samples; error bars represent standard deviation. The extent of lipidation in b and c is plotted as a percentage of total GL1 as determined by densitometry. P values represent comparison with most active liposomes (sonicated DOPE). **P 0.1; *P 0.05.
G. T84 MX-1 mcherry were infected with SARS-CoV-2 at an MOI=3 or MOI=1. 24 hpi cells were incubated in the presence or absence of 2000 IU/mL of IFNβ1. 12 h post-treatment cells were fixed and stained for SARS-CoV-2 N protein. The number of MX-1 positive, SARS-CoV-2 positive and double positive cells was quantified. N=3 biological replicates were performed. Error bar indicates standard deviation.
A-D Cholesterol metabolic parameters were assessed following 5 days of treatment of DIO male mice with vehicle (white), liraglutide (10 nmol/kg) (gray), RM-493 (3.6 μmol/kg) (black), or liraglutide (10 nmol/kg) and RM-493 (3.6 μmol/kg) (checkered). (C) expression of genes implicated in hepatic cholesterol and lipoprotein uptake. Data represent means ± SEM;n = 8; *P < 0.05, **P < 0.01, ***P < 0.001.
B. Heatmap showing the codon expression bias across the transcriptome for different developmental time points (2, 6, 12, 24 and 48 hpf) and adult organs in zebrafish, and Xenopus at 48h (right). The codon optimality, and CSC calculated during embryogenesis is shown to the right (red, optimal to blue, non-optimal). Heatmap shows the correlation of codon occurrence and Xenopus mRNA expression at 48 h. On the left a heat map (Red-Blue) shows the CSC values for zebrafish and Xenopus. The codons are sorted by their CSC values. For most tissues, optimal codons are correlated with highly expressed genes (orange), while non-optimal codons are anticorrelated with high levels of expression (purple). We also observe a correlation between mRNA expression and the CSC across different samples (2h P= 1e-07 R=0.6, 6h P= 2e-16 R=0.7, 12h P= 3e-09 R=0.6, 24h P= 2e-07 R=0.6, 48h P= 1e-07 R=0.6, Brain P= 1e-04 R=0.4, TestisP= 1e-07 R=0.6, Liver P= 1e-03 R=0.4, Spearman rank test).
(C) FYCO1990-1,233 strongly colocalizes with Rab7, LysoTracker red, and Alexa Fluor 647-dextran (10,000 D) but only weakly with PXp40phox. HeLa cells transfected with the indicated constructs or labeled with LysoTracker red for 60 min and Alexa Fluor 647-dextran (10,000 D) for 4 h were imaged 24 h after transfection.
Confirmation that Pnp1 is the physiological purine nucleoside phosphorylase, Urh1 is the pyrimidine hydrolase, and Prm15 (Pgm3) is the phosphoribomutase. Ratio of metabolite levels in pnp1/urh1, pnp1, urh1 and prm15 (pgm3) strains versus wild‐type strain during carbon starvation. Data are shown in heat map format, with each line reflecting the dynamics of the ratio of the metabolite levels in a particular strain versus wild‐type strain. All reported values are log2 transformed ratios; data are mean of duplicate samples at each time point.
(B) Representative images of TH and netrin-1 stainings by immunofluorescence (scale bar, 2000 μm) six weeks after the intranigral injection of CTL adenovirus vector (left panel) or Cre adenovirus vector (right panel), in the midbrain (substantia nigra (SN) and ventral tegmental area (VTA) of Netrin-1fl/fl mice.
C: Image of a NTH/GFP sporozoite, showing NTH::GFP localisation in a tubular structure. Scale bar = 10μm.
Top: 1H spectrum of DFL, where the numbered peaks correspond to DFL protons (left). Middle: superimposition of STD spectra obtained for 0.5 mM DFL with 0.05 mM CXCL12 (black line) and upon addition of 0.2 mM of CXCR41-38sY21 (red line). Bottom: waterLOGSY spectra obtained for 0.5 mM DFL with 0.05 mM CXCL12 (black line) upon addition of 0.2 mM of CXCR41-38sY21 (red line).
(F) Maturation state (CD62L-CD44- double negative, CD62L-CD44+ effector memory, CD62L+CD44- naive and CD62L+CD44+ central memory) of CD8+ lymphocytes (mean ± SD, each dot represents an individual mouse; WT IFNγ vs WT CTRL: **** p ≤ 0.0001, ordinary One-Way ANOVA).
PTX3, released from astrocytes, binds to hyaluronic acid through TSG6 and activates β1-integrin. β1-integrin controls AMPAR trafficking through downstream MAP Kinase activation. Indeed the PTX3-induced AMPAR enhancement is prevented by 1) genetic removal of the key HA-binding factor, TSG6; 2) neuron exposure to hyaluronidase (HAse, which destroys the extracellular matrix; 3) Inhibition of β1-integrin activation; 4) pharmacological blockade of MAPK activation (PD98059)
H) Plaque numbers were measured for Pru (n=12), PruΔgra15 (n=12) and GRA15 complemented parasites (n=6).
O, P. GTT data for MI-463 or vehicle treated PbkKI/KI or PbkWT/WT mice (n=3 for each group) on 8 weeks HFD (O) (*P = 0.0133 (Two way ANOVA), ns, not statistically significant difference.) and AUC of GTT data (P) (*P = 0.0442 (two-tailed unpaired student's t-test), ns, not statistically significant difference.).
and immunoblots for caspase-1, caspase-11, IL-1β, IL-18 and IL-1α (b) from unprimed BMDMs infected for 16h with the indicated bacteria (grown to stationary phase).
LINC00115 regulated ZNF596/EZH2/STAT3 signaling. 1123 and 528 GSCs were treated with or without EZH2 inhibitor GSK343 (3 μM) for 6 Data information: data are representative of three independent experiments. Error bars, ± SD.
Pairwise comparison of H3K27 acetylation in WT and DKO AMs performed on 2-kb regions around the Bhlhe40 peak summits (right - for all Bhlhe40 peaks called with P value of < 10−10; center - for top 3,000 Bhlhe40 peaks (ranked by P value)) and around WT AM ATAC-seq peaks (Immgen) that did not have a Bhlhe40 peak in their close proximity (filtered as described in Materials and Methods) (left). Volcano plots show adjusted P values (vertical axes) and log2 fold change (horizontal axes) for H3K27ac pairwise comparison in WT vs DKO AMs; genes assigned to some of the Bhlhe40-binding regions are labeled on the left plot.
B mDA neurons in the cultures transduced with Nurr1 and/or Foxa2 are resistant to toxic stimuli induced by H2O2 (50-500 μM), 6-OHDA (50-1,000 μM), or MPP+ (50-500 μM). The mDA neuron-glia cultures were transduced with lentiviruses expressing Nurr1 (N), Foxa2 (F), Nurr1 + Foxa2 (NF), or control (C) and treated with the toxin for 8 h. Viable TH+ cells were counted on the following day. Shown in the graphs are percent TH+ cells relative to the respective toxin-untreated cultures. TH+ cells of C, N, F, and NF were compared at the same concentrations of the toxins. Significantly different from the control (C)*, from N#, and from F‡ at P < 0.05, n = 10 cultures each; one-way ANOVA followed by Bonferroni post hoc test.
(F) Tyrosine phosphorylation of EGFR in HBMEC in response to GBS strain K79 infection at 15 min and 60 min post infection.
F. Neuro-like sphere formation assay of effects of re-expression of shRNA-resistant LINC00115 WT and mutant with or without miR-200b. Data information: data are representative of three independent experiments. Error bars, ± SD. **, p < 0.01, by two-tailed t-test in F.
(G) GFP mRNA expression levels were measured by qPCR. RNA levels were normalized to GAPDH, β-actin and β2-microglobulin mRNA. Bar-graphs of mean ± SD. Unpaired two-tailed t-test with Welch's correction. * denotes p<0.05, and ** denotes p<0.01.
The average number of mitophagosomes per section (320 μm × 320 μm) was quantified from the total number of brain slice sections (n) indicated on the top of the bars
A | Intracellular expression of IgM (Ig-μHC ic) in bone marrow (left) and spleen cells (right) was determined by flow cytometry and compared between the populations of B cells (green, identified by B220 and CD19 expression) and non-B cells (gray). Numbers in the histograms indicate the percentages of the positive populations. Figures are representative of 11 - 35 individual mice per genotype.
In the apo state of Sec complex, the Sec61 channel is closed by the plug while the lateral gate is already open. Sec62 flexibly associates with the Sec complex mainly through a basic surface on its Sec62 domain interacting with the acidic C-terminus of Sec63. During the insertion of a post-translational client, its signal sequence (ss) binds to the groove at lateral gate supported by the Sec62 TM2. At the same time, the Sec62 TM1 pushes the Sec61α TM3 outwards leading to an even more open lateral gate. This also leads to removal of the plug away from the pore ring, allowing, the translocating peptide to be gated through the Sec61α central pore.
Transfections were performed as in (A) and cells were stained with an antibody against FG-Nups (mAb414) and Nup133. Representative images for mAb414 staining are shown. Confocal imaging was performed through nuclear cross-sections as well as on the surface of nuclei. (H) To measure NPC densities, confocal NE surface images were acquired for mAb414 and Nup133 stained nuclei. NPC numbers were counted per unit area for 25-116 nuclei per condition (53 nuclei on average) and normalized to the negative control. Two biological replicates, data from one representative experiment shown.
Consensus sequences for KRAB A-Boxes from top to bottom: coelacanth KRAB domains, coelacanth PRDM9 KRAB domain, human PRDM9 KRAB domain and human KRAB domains. Residues conserved between the first three sequences and not present in human KRAB consensus were highlighted in red. In the human KRAB consensus, residues crucial for KAP1 recruitment were highlighted in blue.
Study 4, tumor-bearing male mice receiving AR-42 (10 mg/kg, oral gavage; n=9), TFM-4AS-1 (10 mg/kg, subcutaneous; n=10), Combination of AR-42 and DHT (10 mg/kg oral gavage and 3 mg/kg subcutaneous, respectively; n=10), Combination of AR-42 and TFM-4AS-1 (10 mg/kg, both, n=9) or Vehicle (n=6) and tumor free mice receiving Vehicle (n=6) were treated daily for 12 days starting 6 days after cell injection. A) Terminal body weights corrected for tumor mass according to the Materials and Methods compared to tumor-free controls (mean±SD).
(E) NIH3T3 cells were transfected with the retroviral vectors encoding HA-ULK1K46N or with the corresponding empty retrovirus (mock). They were cultured in complete or starvation medium for 120 min. Cell lysates were then analyzed by immunoblot analysis with the indicated antibodies.
Electron microscopy revealed a number of demyelinated or degenerated axons in PLP-150Q mice at the age of 5 months. LAQ (5 mg/kg) treatment at 3 months of age for 2 months improved myelination in PLP-150Q mice. Scale bars: 2μm (Low magnification) and 0.5 μm (High magnification).
(L) The frequency usage analysis of UUU and UUC codons showed that in 488 genes at least one transcript was found to use the UUU codon for more than 80% of Phe codons. Among the 488 genes, 189 genes are involved in the nervous system and brain functions.
Immunoblot detection of Hydroxylated HIF-1α (HIF-1α-OH), total HIF-1α, Luc, LDHA, BNIP3 and ACTIN levels from normoxic lysates of shCntl and shBNIP3 cells expressing either Luc or HIF-1α-AA and collected 48h after plating (n= 4). Densitometric quantifications relative to ACTIN levels are shown below each corresponding band and they were analyzed with a one-sample T-test against shCntlLuc except for LDHA in shBNIP3 Luc, which was analyzed with Wilcoxon rank test.
(A, B) Sustained local 4-AP administration was associated with increased myelin area and close-to-normal Garea-ratio compared to untreated group, as determined by analysis of 40 randomly chosen myelinated axons from sections of 4 nerves for each group. (**: p<0.0001; ANOVA, with post-hoc comparisons using two-tailed unpaired t-test). 4-AP treated mice had a greater proportion of axons for which the associated myelin area was greater than the mean value for uninjured mice (shown in the green line), with inset figure displaying all axons with values above this mean.
Thermostability analysis via nanoDSF. In detergent solution, Zaprinast (250 μM) increased the thermostability of the Mpc1/Mpc3 hetero-complex (black lines) (n=3) but not of Mpc1 (orange lines) (n=3).
Stable isotope tracing mass spectrometry using 13C-labelled serine in cells treated with DMSO and R406. n = 3.
C, D The mRNA levels of selected inflammatory genes were confirmed by qRT-PCR for lean (ND) (C) and obese (HFD) (D) mice (n = 4).
(H) Western blot analysis of lysates from HeLa, TRIM16KO cells and TRIM16KO cells complemented with TRIM16 deletion constructs where cells were treated with MG132 (20 µM, 2h) and the blot is probed with indicated antibodies.
(O) Average expression of ERS DOWN genes from MPH in single-cells from the hepatobiliary lineage. The hepatoblast-to-hepatocyte and hepatoblast-to-cholangiocyte differentiation paths are indicated with arrows.
(L-M) Dorsal trunk cells stained for Serp. In control embryos (L), Serp is seen exclusively in the tracheal lumen. The cells themselves are not visible in these images. In embryos expressing Shrb::GFP under btl-gal4 (M), Serp is also seen inside the cells, usually in association with or surrounded by Shrb::GFP. Boxed regions are magnified in (L'-M') and shown as single confocal planes.
D Methylated DNA immunoprecipitation (MedIP) with antibodies to 5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC) in BL41 B cells sorted and maintained based on distinct exon 5 inclusion levels. qPCR with primers against the CD45 gene body, as indicated.
Genes induced by CALIC are significantly enriched in those upregulated by hnRNP-L. GSEA comparing the genes repressed by CALIC and hnRNP-L knockdown. NES, normalized enrichment score.
A Sec63-mNeon images of cells of the indicated genotypes harboring the inducible system. Cells were treated with 800 nM estradiol for 6 h. Two examples of each phenotypic class are shown.
De-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).
(E) Analogous experiment to (D), comparing histone binding by GST-Spt5 and GST-Spt5N-3A (with F180A E184A V187A mutations of conserved residues).
(B) RLS analysis for WT, nat4∆, pnc1∆ and double mutant strains in BY4741 genetic background. Values in parenthesis (here and hereafter) indicate mean lifespan. Statistical significance (here and hereafter) was determined by one-way ANOVA test: * p ≤ 0.05; ** p ≤ 0.01; *** p ≤ 0.001; **** p ≤ 0.0001. ns = non-significant.