EMBO/sd-character-level-ner huggingface dataset:
D) Michaelis-Menten analysis of USP1 deletion mutants against ubiquitin-rhodamine (Ub-Rho) shows that USP1ΔL1L3 has significantly higher activity compared to WT and other mutants (n=2, biological replicates).
B. Dot plot of the top-three marker genes for each cell type. The dot size represents the percentage of cells expressing the gene; the color represents average expression across the cell type. Fig.S3B shows an extended list of the marker genes.
(D) Overlay of the primer terminus and active site with an undamaged reference structure (2PFO, black). The catalytic aspartic acids are shown in yellow, the primer terminus in cyan and the incoming dATP in magenta. A water molecule (red) and Mg2+ (neon) are occupying the metal A and metal B sites, respectively.
(G, H) Wild-type GSDMD and GSDMDsusti. (G) or mCherry-GSDMD-C (H) were purified from HEK293T cells and incubated with recombinant p20/p10 complex and His-tagged nucleocapsid in a cleavage buffer, followed by immunoblotting with antibodies against the indicated protein tags or antibody against GSDMD C-termimus. Data information: Experiments were repeated for three times with similar results.
A. Imaging of SPBs (Cnm67-RFP) and Spc72-GFP in PHSL1-CDC20 ama1∆ spo13∆ control cells and cells containing cdc5-as or ime2-as treated with CMK or 1Na-PP1, respectively, at metaphase I (7 h in SPM). Top, time-lapse series. Bottom, Spc72's presence at SPBs was quantified in cells synchronized in silico to SPB separation at entry into metaphase I (t = 0). Data information: data are representative of two independent experiments.
D Volcano plot of the result from quantitative transcriptome profiling in WT and dja6 dja5 mutant. Statistical analysis was performed using Student's t-test. Data on the X-axis represent the Log2-ratio of FC (fold change) between the absolute abundances of transcripts identified from dja6 dja5 against WT. The Y-axis represents the Log10 of FDR (false discovery rate). All DEGs (differentially expressed genes) with FDR < 0.01 and FC > 2 are marked in grey, while all iron-related DEGs with FDR < 0.01 and FC > 2 are marked in red. The iron-related DEGs labeled with red dots within the black boxes are referred to in the text. All data are based on three biological replicates.
(E) WT, Irgm2-/-, Casp11-/- and Irgm2-/- Casp11-/- BMMs were treated with OMVs at indicated concentrations for 24 h. Cell viability was assessed via CellTiter-Glo and cell death was calculated as a function of relative viability to untreated cells.
(A) Co‐immunoprecipitation of endogenous NAF‐1 and BCL‐2. Lysates from SK‐Mel5 cells were collected and immunoprecipitation was performed with anti‐NAF‐1 antibody. The precipitate was subjected to immunoblotting with anti‐BCL‐2 and anti‐NAF‐1.
F-K, Growth curves (F, I) of KRAS-mutant lung cancer PDXs (PF139, PF563) treated with vehicle, HCQ (30 mg/kg/day), AZD4547 (10 mg/kg/day) and BI6727 (5 mg/kg/day), alone or in combination. *P<0.05, **P<0.01, ***P<0.001 by one-way ANOVA with Tukey's multiple comparisons test. Data are the mean of tumor volume of each group (4 mice/group); error bar: SD Relative tumor volume (G, J) and weights (H, K) of the PDX tumors after the treatment. *P<0.05, **P<0.01, ***P<0.001 and P＞0.05 (ns) by unpaired two-tailed t-test. Data are the mean ± SD (error bar) of tumor weights of each group (4 mice/group).
(A) INS1 cells were transduced with mtPA‐GFP (green) and stained with TMRE (red). mtPA‐GFP distribution is shown immediately (within 2 min) after photoactivation (area indicated by a dashed square) and in the plateau phase of mtPA‐GFP spreading (45 min). Mitochondria that did not exhibit GFP FI decay (less than 5%) are marked by arrows. Upper scale bar, 10 μm; lower scale bar 5 μm.
(a-k) Confocal analyses of the subcellular co-localization of endogenous UVRAG with ER markers, including endogenous calnexin (red, a), endogenous PDI (red, b), and overexpressed DSred-ER (red, c); with Golgi markers, including endogenous GM130 and p115 (red, d,e, cis-Golgi), overexpressed GFP-Man II (green, f, medial-Golgi), endogenous TGN46 (red, g, trans-Golgi), and overexpressed PtdIns(4) P probe, GFP-FAPP1-PH (green, h); with endogenous coatomer proteins, including Sec31 (red, i, COPII-related) and β′-COP (red, j, COPI-related); and with the endosome marker GFP-p40phox(PX) (green, k) in HeLa cells. The insets show a high magnification of the selected areas. Scale bars, 10 μm. (l) Confocal microscopic quantification of co-localization of UVRAG with the indicated markers (data are mean±s.d., n = 150 cells obtained by gathering data from three independent experiments); **P0.01; ***P0.001.
Workflow of the SILAC-based quantitative mass spectrometry method for identifying Dpb11 interactions.
D The SARS-CoV-2 RBD from A is shown as a ribbon, colored according to the indicated color gradient, with its N-terminus in blue and C-terminus in red. Disulfide bridges are depicted as yellow sticks. Re9F06 has been omitted for clarity. RBD side chains that interact with the fold-promoting VHH Re9F06 are shown as green "ball-and-sticks". See Appendix Fig S5 for a comprehensive analysis of the Re9F06⋅RBD interface.
Alkaline Phosphatase (ALP) levels in the serum at endpoint (age = 5-6 months; n=16 wa/+, 11 wa/wa)
SSA/RMD and BIR/SSA are both dependent on ATM. SSA/RMD and BIR/SSA frequencies in U2OS (EGFP-SSA-3427-17) cells were determined after the addition of ATM inhibitor (20mM ku55933) or DMSO to the cell culture medium 24 hours before transfection of the indicated sgRNA/Cas9
Genomic location and sequence of a putative AP-1 binding site in the mouse promoter of IL-23p19. Blockade of c-Jun/AP-1 TF binding by inhibition of phosphorylation (JNKi, red), or mutation of the binding sequence (mut 1 = T -> C; green | mut 2 = TG -> AT; blue) reduces IL-23p19 promoter activity (see Fig 4J).
Representative images of microtubules (top) and actin filaments (bottom) growth from micropatterns. Scale bars: 20 μm.
(K) A schematic diagram depicting the regulation of lipogenesis by PKM2. ER located PKM2 inhibits VCP activity, which is essential for NRF1 precursor cleavage. Cleaved NRF1 acts as a transcriptional factor for TMEM33. ER localized TMEM33 inhibits SREBPs activity through promoting SCAP degradation in RNF5-dependent manner. Knocking out PKM2 leads to increased NRF1 cleavage and TMEM33 expression. Consequentially, SCAP is destabilized by RNF5-mediated ubiquitination and expression levels of lipid synthetic genes are decreased.
(H) The number of cells with intact Golgi was divided by the number of total cells per image and plotted against time (three biological replicates). Dots are means ±SD. For quantification, a sigmoidal fit was performed with the software GraphPad Prism 7. Dashed lines indicate extrapolated plateaus. LogIC50 ±95% confidence interval: C1 = 59.38±2.31 min, P1 = 72.89±1.65 min, P2 = 68.57±1.929 min. Significant differences were assumed when 95% confidence interval do not overlap.
B: Signalling input conditions (left side shows deduced input function, right side shows pERK2 measurements): Sustained ERK signalling (4OHT), two-hour pulse ERK signalling (4OHT+U0126), growth factor signalling (EGF: Epidermal growth factor, FGF: Fibroblast growth factor). Deduced input functions: 100% signalling amplitude corresponds to mean induction in training condition (4OHT). Growth factor-induced input functions are linear interpolations of pERK2 log2 fold changes relative to mean induction in test condition.
(L) Representative FACS plots showing the percentage of SCs sorted from 6-month-old Ctrl and YY1dKO mice (n=3 mice, each).
C. Endogenous co-IP of USP44 and FOXP3 in murine nTregs. nTregs (CD4+CD25high) isolated by FACS were activated by anti-CD3 and anti-CD28 (1 and 4 ug/ml, respectively) overnight in the presence of IL-2 (100 U/ml). The cells were lysed and proteins were immunoprecipitated using either anti-Foxp3 (left panel) or anti-Usp44 (right panel). Proteins pulled down in this experiment were then resolved and identified with the indicated antibodies.
C. SecA ATPase activity determination in solution (basal; B), or plus SecYEG IMVs; (membrane; M) or plus proOmpA (pre-treated with 10 mM DTT; 5 mM EDTA; 20 min; 4°C; translocation; T). n= 21 biological replicates. Mean values (± SEM) shown.
D. Inducible mepS expression system (pBAD30) strongly overproduces MepS. Strains were grown in LB at 30°C and cells were collected at OD600≈0.4. The level of MepS contained in the membrane fraction was detected using purified anti-MepS antibody.
A. Schematic drawing of 2-step QTIP (improved version of the quantitative telomeric chromatin isolation protocol (QTIP)).
g, Quantitative effects of beclin 1 on basal and nutrient-deprivation-induced autophagy of MCF7 cells. Bars indicate mean (±s.e.m.) number of autophagic vacuoles per cell for cells growing in normal media (black), for cells subjected to 4 h of serum and amino-acid deprivation (grey), and for cells pre-treated for 30 min with 10 mM 3-MA and subjected to 4 h of serum and amino-acid deprivation in the presence of 3-MA (open). Mean was determined by counting the total number of autophagic vacuoles in each cell for 100 cells per clone per treatment.
(e) Interaction between endogenous UVRAG and the RINT-1-ZW10-NAG tethering complex proteins. WCLs of HEK293T cells were used for immunoprecipitation with control serum (IgG) or an anti-UVRAG (first panel), anti-RINT-1 (second panel), anti-ZW10 (third panel) and anti-NAG (fourth panel), followed by immunoblotting with the indicated antibodies. As input, 10% of the WCLs was used. Syn 18, syntaxin 18; Syn 6, syntaxin 6. See Supplementary Fig. S9 for uncropped data.
C. Average database release cycle length for releases between August 2013 - July 2019. All values are mean ± SD. Total of 25 Ensembl, 13 GENCODE, 6 CCDS (homo sapien only), and 5 NCBI releases were counted. UniProtKB value was calculated by taking the average of release cycle lengths reported on the UniProt website.
Localization of SPATC1L and RIα during spermatogenesis. Isolated spermatogenic cells were immunostained for SPATC1L (red) and RIα (green), and counterstained with the nuclear dye Hoechst 33342 (blue). Overlay with bright field image is shown at right in each cell type. TC, testicular cell (spermatocyte); RS, round spermatid; ES, elongating spermatid; TS, testicular sperm. Scale bar = 5 μm.
(F) Proliferation of WT and Mettl14 null cells in vitro was quantified and shown as dot plots. n=3, P<0.01 (Student's t-test) for Day 6, 9, and 12, and P<0.05 (Student's t-test) for Day 3. Error bar represents S.D.
Representative cryo-EM image where roughly half of the particles correspond to 40S (blue) and the other half to 80S (orange).
Radiolabeling with [35S]methionine, anti-FLAG immunoisolation and sample analyses were performed Total 1 %, Eluates 100 %. Red asterisks, accumulating nascent chains of ND2.
A, Intravitreal injection of LIF increases blood vessel density in the mouse eyes. Adult mice were intravitreally injected with VEGF (10 ng) or LIF (10 ng to 100 ng, Sigma). Seven days after injection, PFA-fixed choroid-sclera complexes and retina were subjected to CD31 IF. Representative images of CD31-positive vessels are shown in A. Scale bar = 100 µm.Vascular
A Ensemble FRET assay showing the different effects of the Munc18-1 P335A mutation and the Syx1 LEAA mutation in the transition from the Munc18-1/Syx1 complex to the SNARE complex with and without the MUN domain. FRET signal between BODIPY FL-labeled Syb2 (donor) and TMR-labeled SN25 (acceptor) was monitored.
E) Whole cell expression of PAR in mBA (WT: Ews-WT, KO: Ews-KO, DKO: double KO).
(F) Western blot and densitometry analysis of duodenal zonulin (n = 6). Data information: Data were representative and were the mean ± SD from three independent experiments. P values were calculated by one-way ANOVA followed by Tukey's post hoc test for multiple comparisons.
After 2 hour starvation, E. coli cultures were pulse fed 10 µM glucose at varying frequencies using the spin flask and plate reader systems, and optical density (OD) was measured over time (inset example figures). Grey dots are OD measurements and the black lines are an empirical fit (see Methods). For separate experiments (n = 18), the lag time is plotted against the frequency, represented as the time-integrated (TI) feedrate f (mmol glucose/g dry cell weight/hour). An empirical fit (grey solid line, see Methods) was used to separate the lag (non-dividing) and dividing phases
C. The plots represent the discrimination index measured in each condition. *** p < 0.001, one-way ANOVA followed by Bonferroni test. The discrimination index for the novel object recognition task is 0.34 ± 0.06 (N=14) and -0.08 ± 0.07 (N= 20), for mice injected with Vehicle and THC, respectively, p < 0.001 and -0.09 ± 0.08 (N=21) and 0.00 ± 0.06 (N=18), for mice treated with THC+SB258285 and THC+Rapa, respectively, p > 0.05 vs. THC-injected mice (error bars correspond to the mean ± SEM, the dotted line to a discrimination index equal to zero).
Figure 1. Extensive identification of Mmi1 RNA targets by combining RNA-IP sequencing and computational approaches. (C) Surface representation of Mmi1 YTH domain. Area corresponding to conserved residues (according to the alignment shown in Fig EV 2A) and located at the surface are highlighted in green. Mutated residues in Mmi1 YTH domain are indicated in blue for the ones located within the aromatic cage, in purple for the ones surrounding the cage, and in red for the rest.
B RT-PCR analysis with the indicated primers in A for control and Gr28bMi flies with tubulin as a control. From left to right, the band of tubulin of wild-type, tubulin of Gr28bMi, DNA ladder, Gr28b of wild-type, and Gr28b of Gr28bMi.
Matrigel invasion assay of MDA231-LM2 cells treated with JNKi. Left, examples of invading cancer cells stained with crystal violet (arrows). Scale bar, 50 μm. Right, quantification of invading cells per field of view (FOV) showing mean from 8 fields ± SEM.
(A) Body length of individual nsun mutants throughout development (n = 44,7,7,7,8,8) in ~4 hour windows. L1-L4 refers to the larval stages, YA and Ad to young adult and adult, respectively. Data information: In (A), error bars indicate the 95% confidence interval of the median.
C: The E14tg2a ESCs were transduced with lentiviral shRNAs targeting genes Atp5g3, Bub1b, Eci1, or Dis3l and induced to form EBs. The day 6 EB cells were analyzed by flow cytometry to measure the Mesoderm-Endothelium cells (Flk1+/Cxcr4-) and Endoderm cells (Cxcr4+/Flk1-). Flow cytometric analysis showed that Endoderm cells (Cxcr4+/Flk1-) were deceased while Mesoderm-Endothelium cells (Flk1+/Cxcr4-) were slightly increased in the cells with shRNA knockdown of Group XI genes Atp5g3, Bub1b, Eci1, and Dis3l compared to control.D: Knockdown of Atp5g3, Bub1b, Eci1, or Dis3l led to increase of Mesoderm-Endothelium/Endoderm population ratio as quantified based on the flow cytometry data. Error bars represent standard deviation (SD), n=3. Student's t-test, *p<0.05.
B) A 39 bp fragment of ub-HS4 is sufficient to drive ectopic ub-HS formation. Fragments of ub-HS4 were integrated ectopically and ub-K48 ChIP-qPCR was performed for the endogenous ub-HS5 and the ectopic ub-HS4 fragments as depicted in (A). contr.: control, empty YI128-vector was integrated at LEU2. Experiments were performed in cdc48‑6 background. Data represent means ± SD (n = 2). Experimental mapping and bioinformatic prediction identify a similar ub-HS-motif. Comparison between the experimentally mapped ub-HS4 F7 (B) and the consensus motif of all ub-HSs identified by the MEME software.
quantification (D) of L-homopropargylglycine (HPG) showing the overall translation levels of MI and MII oocytes collected from WT and Cnot6l-/- mice. Scale bar, 20 μm. Error bars, standard deviations (n = 6 oocytes for each genotype). **P < 0.01, ***P < 0.001
(A) Quantification of AQR position in mig-2(mu28) null mutant animals with different rescue transgenes. N = 108 - 114. *, P < 0.05; ##, P < 0.01, phenotype enhanced. P values are based on Fisher's exact tests.
A. Kinase activity in yeast. Kinase activity is played on the human kinome tree showing that all kinase families contain highly active kinases members.
A. Representative image of a Drosophila posterior midgut showing the expression of Armadillo (red membranes), Prospero (red nuclei of EEs) and Dronc-TurboID (green shows Streptavidin labelled proteins by Dronc-TurboID); note the preferential enrichment of Streptavidin staining is preferentially enriched in the intestinal progenitor cells (white arrows indicate an intestinal progenitor cell upregulating Dronc).
(E) Venn diagram shows the number of GFP RNA foci, the number of LUC RNA foci, and the number of GFP RNA foci colocalizing with LUC RNA foci.
(C) GFP-Atg8 vesicles colocalized with glycogen in Dmef2-Gal4, UAS-GFP-Atg8 larvae starved on low-nutrient food +2.5 mg/ml CQ for 6 h (GFP, green; antiglycogen, red).
Synaptic transmission in autaptic excitatory munc18-1 neurons expressing M18WT or a non-phosphorylatable Munc18-1 variant, M18Y473F, was tested with whole-cell patch clamp electrophysiology. (A) Evoked release upon action potential stimulation. Average EPSC amplitude (M18WT: 3.78 ± 0.58 nA, n=22; M18Y473F: 4.10 ± 0.61 nA, n=20, Mann-Whitney U test, p = 0.7915). Typical responses are depicted on the top.
(B) RIBO-seq coverage profiles from 35S:EVD in WT or rdr6. RPM: Reads per million. (C) Ribosomal footprints on shGAG in rdr6 displaying codon occupancy at P-sites to calculate codon coverage. The coverage observed at each codon position was divided by the expected mean coverage along the entire GAG coding sequence.
(A) SARS-CoV-2 maximum likelihood phylogenetic tree. The SARS-CoV-2 isolates used in this study are depicted with black lines and labeled. The continents where the virus strains were from are colored differently.
(A-D) Time course of autophagy induction in Dmef2-Gal4, UAS-GFP-Atg8 muscles, accompanied by quantification of GFP-Atg8 and glycogen colocalization. Animals were fed for 18 h in high-nutrient food +2.5 mg/ml CQ, then starved on low-nutrient food +2.5 mg/ml CQ for 0-8 h (antiglycogen, red; GFP, green; DAPI, blue). (A) At time point 0, following 18 h in high-nutrient food +CQ, the muscles contained large amounts of glycogen with no apparent autophagy. (B) At 3 h of starvation, glycogen stores were still high, and GFP-Atg8-labeled vesicles began to appear. (C-D) At 6 and 8 h of starvation, the majority of GFP-Atg8-labeled vesicles colocalized with glycogen.
i to o. Inhibitory activity of EK1C4 in PsV infection assays against SARS-CoV (i)
A-D. Mice were injected daily with THC (5 mg/kg) or vehicle (Veh) during adolescence, from PND 30 to 45. SB258585 (SB, 2.5 mg/kg) or rapamycin (Rapa, 1.5 mg/kg) were administered concomitantly with THC (n=20 from N=5) or vehicle (n=18 from N=6). Electrophysiological recordings were performed from PND 60. D. The firing rate induced by a 150 pA current injection during 250 ms is expressed as the number of APs.
The interaction of endogenous TRAF2, ASS1 and LRPPRC was detected in SW-1116 cells by co-IP assays. GST-TRAF2, HA-LRPPRC, His-ASS1 and Biotin-LOC113230 were expressed in E. coli BL21 cells, and were used for in vitro binding assays.
(B) Venn diagram shows the number of vRNA clusters, the number of vDNA clusters, and the number of vRNA clusters colocalizing with vDNA clusters. The p-value indicates the significance of colocalization based on a jittering analysis (see Methods).
(D) Lipid mixing was monitored by the increase of Atto 655 fluorescence for 30 minutes. Data were normalized to the maximum fluorescence after liposome lysis by detergent. The minimum fluorescence was set to 0%. Error bars = s.e.m., N = 3.
(D) Quantification of CTxB transport to the Golgi apparatus in BMMs following siRNA-mediated depletion of either Arf6 (siArf6), Rab8a (siRab8a) or Rab6a/a' (siRab6a/a') after AlexaFluor™488-CTxB binding on ice followed by 30 min incubation at 37˚C. Data are means ± SD from n=3 independent experiments, in which 100 cells were analyzed per experiment. Asterisks indicate a statistically significant difference compared to siNT control cells as determined by a one-way ANOVA with Tukey's multiple comparisons test (P<0.05). (E) Quantification of CTxB transport to the Golgi apparatus in BMMs that were either mock-infected or infected with wild type (2308), ΔbspF, complemented ∆bspF (ΔbspF::bspF), ∆bspB or complemented ∆bspB (ΔbspB::bspB) bacteria for 24 h, incubated for 30 min with AlexaFluor™488-CTxB on ice for binding followed by 30 min incubation at 37˚C. Data are means ± SD from n=3 independent experiments, in which 100 cells were analyzed per experiment. Asterisks indicate a statistically significant difference compared to mock-infected cells as determined by a one-way ANOVA with Tukey's multiple comparisons test (P<0.05). (
RT-PCR analysis in P30 mice showing the presence of Clrn2 transcripts in the inner ear, eye, but not in brain or muscle. β-actin was used as a positive control.
D. Upregulated gene numbers based on different expression level in control and MOSAsn95Lys MII oocytes.
B. A549 cells infected with ZIKV (MOI=3) for 6 hrs were transfected with 2 μg poly(I:C) for 12 hrs after which levels of ifn-
a, CD loop folding in hydrophobic buffer. Peptides were synthesized that correspond to the predicted amphipathic helix from S. cerevisiae Atg18 (TRLAREPYVDASRKTMGRMIRYSSQ) or to a sequence with reduced tendency to form amphipathic helices, generated by a swap of two amino acid pairs (indicated by arrows; SLoop). Helical wheel projections of the sequences are shown.
Immunoblot (left) and densitometry (right) analysis of IκBα levels in primary healthy control and patient fibroblasts treated with cycloheximide (CHX) (50 μg/mL) as indicated. Data are representative of three independent experiments.
Relative average expression levels of IL1B and NLRP3 in non-classical monocytes of each group were calculated (C). n=20 for healthy donor, n=9 for mild/moderate, n=7 for severe. each dot represents an individual. Brown-Forsythe and Welch ANOVA test was used and data were shown as means±SD. *, P<0.05; **, P<0.01; NS, non-significant. Data information: Brown-Forsythe and Welch ANOVA test was used and data were shown as means±SD.
qRT-PCR analysis of FOXP2, FOXC2 and GJA4 expression in control (siCTRL) and FOXP2 or FOXC2 siRNA-treated HDLECs (n = 6 (FOXP2, FOXC2) or n = 3 (GJA4) independent experiments).
B Cell duplication rate of primary hepatocytes from P240 PR-SET7ΔHepA livers in culture.
C) Graph showing significantly (p = 0.029 [**]) increased K15/caspase 3 double+ cells within the bulge following extended paclitaxel HF organ cultures. Welch's t-test performed using N of 8-9 HFs from 3 patients. Values plotted represent the mean number of positive cells counted per HF analysed. Error bars are standard error.
C) Tmem106b-/-Grn-/- mice show reduced activities in the open field test. 4.5-month-old mice were used in the study. Total movement was quantified. n=8-10. Data presented as mean ± SEM. One-way ANOVA tests with Bonferroni's multiple comparisons: ****, p<0.0001.
a, Immunoblot analysis of Atg5 and LC3. Brain homogenates were prepared from six-week-old control (Atg5flox/+; nestin-Cre) and Atg5flox/flox; nestin-Cre (Atg5flox/flox) mice. Immunoblot analysis was performed using antibodies against Atg5 and LC3. GSK-3β was used as a loading control. The positions of the Atg12-Atg5 conjugate, LC3-I and LC3-II (LC3-PE conjugate) are indicated. Atg5 monomer was not detected in either lane (data not shown).
(E) Control and FRS2 knockdown HASMCs were cultured in the growth medium (M231 + SMGS) at day 0 then switched from growth conditions to differentiation medium (M231 + SMDS) for 6 days with or without let-7b lentiviruses. Immunoblots of smooth muscle markers, phosphorylated Smad2 (p-Smad2), and TGFR1 expression in control and FRS2 knockdown HASMCs with or without let-7b lentiviruses. Blots are representative of three independent experiments.