EMBO/sd-character-level-ner huggingface dataset:
Figure 2A. Sequence of RNA substrate used for in vitro exoribonuclease assays. N, randomized nucleotide. B. In vitro exoribonuclease assay using 5′ radiolabeled RNA substrate and immunopurified wild-type (WT) or catalytic mutant (CM) FLAG-dmDis3l2 (mutation indicated in Figure 1C), incubated for the indicated time and separated on a 15 % polyacrylamide gel followed by phosphorimaging. Quantification of degraded substrate in percent is indicated.C. Change in abundance of 256 different substrate RNAs as determined by high-throughput sequencing of substrates in experiment shown in (B). D. Decay rates of 256 different substrate RNAs. Data shown in (C) was normalized to overall decrease in substrate abundance as determined by phosphorimaging (as shown in B) and fit to the indicated model for exponential decay. Apparent decay rate of all substrates, as determined by phosphorimaging is indicated in red.E. Selected examples for individual substrates shown in (D). Values report the observed decay rate (kobs) in min-1.F. Overview of decay rates (kobs) of all 256 different substrate RNAs, as determined in (D). Error represents SEM of curve fit. G. Impact of RNA secondary structures, as determined by RNAfold (Gruber et al, 2008), and reported as effective free energy (EFE, kcal/mol), on decay rates (kobs) are shown. Error bars represent SEM of curve fit. Grey boxes represent tukey boxplot of individual EFE groups. White line indicates median. P-Value was determined by Mann-Whitney test comparing individual EFE groups against all substrates. Unstructured substrates used for subsequent analysis are indicated (EFE groups 4 to 6). H. Nucleotide content of single-stranded RNA substrates, ranked from slow (top) to fast (bottom) decay by dmDis3l2.I. Cumulative distribution of decay rates for RNA substrates with the indicated nucleotide at each one of four randomized position at the 3′ end of the substrate RNA. P-Value was determined by Mann-Whitney test comparing individual nts against all. J. Functional coupling between two base positions within the randomized 3′ end of substrate RNA. Red squares show average increase in decay rate compared to all substrates. Black squares show no or negative effect. K. Boxplot decay rates of substrates containing no, one, two, or three uridine(s) within the randomized 3′ end of the substrate RNA. Median decay rates SEM of fit are indicated.L. Decay rate of all substrates, or substrates containing no, one, two, or three uridine(s) within the randomized 3′ end of the substrate RNA. Median and quartile ranges are indicated. P-values were determined by Mann-Whitney test. Substrate containing 4 uridines (4U) is indicated.
Figure 6PLXNB2 expressions in HCT116 (including wild type HCT116 (HCT116WT) and ANG knockout HCT116 (HCT116KO)) and THP-1 as detected by flow cytometry analysis. The upper three dash lines represent the intensity of unstained negative control of these cells, while the solid lines represent the intensity of PLXNB2 antibody staining. Secreted ANG level in conditioned medium from THP1, HCT116WT or HCT116KO (n = 3). The interaction between ANGWT, ANGR66A or ANGK40I with PLXNB2 in HCT116KO detected by co-immunoprecipitation assay. ANGR66A, ANG receptor binding site variant; ANGK40I, ANG enzymatic activity null variant THP-1-derived ANG translocating into HCT116KO through PLXNB2 in co-cultured system as showed by western blotting. Neamine, a blocker of ANG and PLXNB2 interaction; siCtrl: control siRNA; siPLXNB2-1 and siPLXNB2-2 Flow cytometry analysis of HCT116KO stained with Annexin V and PI following different treatments as indicated. ANGR66A, ANG receptor binding site variant; ANGK40I, ANG enzymatic activity null variant; BAY, inhibitor of TNF-α-induced κB kinase (IKK) phosphorylation tiRNA and 5'-tiRNAAla production in HCT116KO following different treatments as indicated. ANGR66A, ANG receptor binding site variant; ANGK40I, ANG enzymatic activity null variant; BAY, inhibitor of TNF-α-induced κB kinase (IKK) phosphorylation The effect of 5'-tiRNA knockdown or overexpression on TNF-α-induced apoptosis in ANGWT- or ANGK40I-treated HCT116KO. si5'-tiRNA: tiRNA-targeting siRNA The interaction between 5'-tiRNA and cytochrome c (Cyt c) in HCT116 cells as detected by RIP. Caspase activity of HCT116KO following different treatments as indicated. ANGR66A, ANG receptor binding site variant; ANGK40I, ANG enzymatic activity null variant; BAY, inhibitor of TNF-α-induced κB kinase (IKK) phosphorylation Cell viability of HCT116KO following different treatments as indicated. siCtrl: control siRNA; siPLXNB2-1 and siPLXNB2-2: PLXNB2 siRNAs Detection of 47S pre-rRNA transcription level with northern blotting in HCT116KO following different treatments as indicated. siPLXNB2-1 and siPLXNB2-2: PLXNB2 siRNAs tiRNA and 5'-tiRNAAla production in IECs isolated from WT and Ang-/- colitis mice at indicated time point. 47S pre-rRNA transcription level in IECs from WT and Ang-/- colitis mice at indicated time point.
Figure 5Construction of the network EDNRB (green diamond) was connected to its target phosphorylation sites (red hexagons) through intermediary kinases or G-Proteins (blue circles) in a time resolved variant of the PHONEMeS approach. Central kinases, which were also identified by kinase activation prediction are shown as intermediary kinases with dark blue shading. Edge thickness corresponds to weights, which were assigned by downsampling the network 100 times. Entry time point was defined as the point at which edge weight reached 20 and is shown as edge colour. The network was divided into five modules, indicated as light blue outlines and labelled 1-5. Common names are shown for kinases and primary gene names are shown for all other proteins.
Aggresome‐targeted proteins are degraded by autophagy.Figure 3(A) Analysis of SODG85R‐GFP (green) localization with aggresome marker vimentin, the autophagic receptors p62 and NBR1, the autophagosome marker LC3 and the lysosomal marker LAMP2 (all in red) in BAG3/Hsp70‐transfected HEK cells. Representative merged images of cells with cytoplasmic, pre‐aggresomal and aggresomal SOD distribution are shown. Arrows indicate LC3‐positive autophagosomes. Scale bar, 5 μm. (B) Transmission electron microscope analysis of SODG85R‐GFP aggresome in cells transfected as in (A). Marked area in (a) is shown magnified in (b). Arrows, autophagosomes. Arrowheads, autolysosomes. Scale bar, 1 μm. (C) HEK cells transfected for 8 h with SODG85R‐GFP along with FLAG‐BAG3 and HA‐Hsp70, as indicated, were treated for 24 h with BafA1 (0.1 μM). Indicated proteins were analysed by immunoblotting. Diagram shows the ratio of SODG85R‐GFP levels in BafA1‐treated cells to those in untreated cells. (D) Same analysis as in (C) but with SODWT‐GFP. (E) Similar analysis as in (C) but in cells transfected 24 h before with nonsense or atg7 siRNA. (F) HEK cells were transfected with SODG85R‐GFP and indicated FLAG‐tagged BAG3 constructs together with HA‐Hsp70, and lysosomal degradation of SODG85R‐GFP was examined as in (C). Corresponding immunoblots are shown in supplementary Fig S3F online. Ag, aggresome; BafA1, bafilomycin A1; GFP, green fluorescent protein; HA, haemagglutinin; Hsp70, heat‐shock protein 70; N, nucleus; NS, not significant; nons, nonsense; PxxP, proline‐rich repeat; siRNA, short interfering RNA; SOD, superoxide dismutase; SODWT, wild‐type SOD.
Figure 6 A schematic illustration of the proposed model depicting ADAR1's involvement in the HIF-1α signalling pathway. ADAR1 mediates RNA editing on two negative regulators of the HIF-1α protein, HIF1A-AS and LIMD1. These events correlate with reduced expression of both factors, albeit via distinct mechanisms. Consequently, ADAR1 contributes to a balanced HIF-1α pool critical for the cellular response to hypoxia. These findings provide a mechanistic explanation to the tumorigenic role of the RNA editor.
Figure 1A Domain architecture of S. pombe bromodomain AAA-ATPases. The D1 and D2 ATPase domains (AAA) are shaded blue and orange, respectively, and the bromodomain (BD) is grey. The percentage similarity to each other and the indicated proteins was determined by FASTA sequence comparison [74] using the scoring matrix BLOSUM50. B Venn diagrams showing overlap between genes up-regulated (1.5 fold) in abo1Δ mutants with genes up-regulated under the indicated condition, along with the significance of the overlaps (based on hypergeometric distribution).
Figure 4The cells specified below were loaded into the upper chamber of a Transwell migration device without (no) or with the specified chemokines in the lower chamber. The percentage of cells that migrated to the bottom chamber after 2 h (T cells) or 4 h (B cells) is shown. A Chemotaxis of naïve CD4+ T cells from WT and Arghap45−/− mice in response to CCL19, CCL21 and CXCL12. B Chemotaxis of naïve CD8+ T cells from WT and Arghap45−/− mice in response to CCL19, CCL21 and CXCL12. C Chemotaxis of naïve B cells from WT and Arghap45−/− mice in response to CCL19 D Adding CCL19 in both the upper and lower chambers (CCL19 UL) prevented migration of WT and Arghap45−/− naïve T cells whereas the use of Transwell membrane inserts precoated with ICAM-1 (CCL19 + ICAM-1) did not change the pattern of chemotaxis observed using 'bare' membrane inserts (CCL19). E Chemotaxis of naive CD4+ T cells and B cells from WT, Arghap45−/−, and Arhgap45∆T/∆T mice in response to CCL19. F Chemotaxis of activated CD4+ T cells from WT and Arghap45−/− mice in response to CCL19. Data information: In (A) to (F) each dot represents an individual mouse and as shown CD4+ T cells from 3 to 9 mice have been analyzed for each condition. Mice analyzed on the same day are represented by similar dots (see key in bottom right corner). Mean and SD are shown. *P ≤ 0.033, ** P ≤ 0.002, *** P ≤ 0.001, **** P ≤ 0.0001; A, B, C, D, F multiple unpaired T test; E two-way ANOVA test.
Figure 4A. Co-immunoprecipitation of Larp with MDI-myc in transfected S2 cells. Tubulin was used as loading control. B. mtDNA replication, as illustrated by EdU incorporation, in wt and larp mutant (larp) ovarioles. mtDNA replication is dramatically reduced in the larp ovariole. Arrows: mtDNA; arrowheads: nuclei. C. wt and mdi1 egg chambers stained for Larp (green) and ATP-S (red) to reveal mitochondria. Larp closely associates with mitochondria in wt egg chambers. Mitochondria in mdi1 flies are clumped together, and completely lack Larp staining. Bars in (B) (C): 10 m.
Micrograph and LC3B antibody labelling of adipocytes neighboring the nerve cells of Porcellio d.Figure 5Micrograph and LC3B antibody labelling of adipocytes neighboring the nerve cells of Porcellio d. dilatatus.These adipocytes are localised around the nerve cells of the CNS and constitutes the sheathing of this organ. (A) At 30 days post-injection, autolysosomes, autophagosomes but also bacteria were observed: autophagic process was already higher compared to the control (Ad: adipocyte, Ap: Autophagosomes (Autophagic hallmark), Al: Autolysosomes, Wo: Wolbachia, Arrow head: magnified area). (B) At 60 days post-injection, the adipocytes were severely damaged by the accumulation of autophagic vesicles (Ad: adipocyte, Ap: Autophagosomes, Al: Autolysosomes, Wo: Wolbachia, Arrow head: magnified area). (C) With the antibody labelling anti-LC3B, the protein LC3B implicated in autophagic processes was detected in adipocytes of P. d. dilatatus injected with wVulC at 45 days post-injection. The highly labeled spherical structures would reflect the incorporation of the LC3B protein in the phagophores. (D) With adipocytes of P. d. dilatatus injected by wDil but also in other tested situations, only really few of these structures were detected by epiflorescent microscopy.
Figure 8a, Schematic representation showing the localisation of SHH components at the cilium in the OFF and ON state(Wheway et al, 2018). See text for details. b-c, Smo translocation to the cilium changes in the presence or absence of Septins. NIH3T3 cells with control, SEPT2 or SEPT7 siRNAs and 48h serum starvation (SS), and then treated with SHH ligands for 0, 1, 2 and 4h. Smo translocation into the cilium is followed over time using the indicated antibodies. Representative images (b) and quantifications (c) are shown. Scale bar: 2µm. Three biological replicates, n>100 cilia per sample and repetition. Data include mean ± s.d. and P values are calculated by two-tailed unpaired student t-test. d, Relative Gli1 mRNA level changes in control and SEPT2 siRNA-treated NIH3T3 cells. Changes in mRNA level are determined by RT-PCR after 0, 4 and 6h of SHH ligand addition to ciliated NIH3T3 cells. Three biological replicates. Data include mean ± s.d. and P values are calculated by two-tailed unpaired student t-test. e, Relative Gli1 mRNA level changes in indicated siRNA-treated NIH3T3 cells. Changes in mRNA level are determined by RT-PCR after 6h of SHH ligand or water addition to ciliated NIH3T3 cells Three biological replicates. Data include mean ± s.d. and P values are calculated by two-tailed unpaired student t-test. Effect of Septins, KIF7 or MKS3 depletion on the localisation of Gli3 and Sufu at the cilia tip. NIH3T3 cells are treated with the indicated siRNAs and 48h SS. Representative images are shown. White arrows in (f) point to the specific signal of the indicated proteins at the cilia tip. Scale bar: 2µm. Effect of Septins, KIF7 or MKS3 depletion on the localisation of Gli3 and Sufu at the cilia tip. NIH3T3 cells are treated with the indicated siRNAs and 48h SS. quantifications of Gli3 and Sufu at the cilia tip (g) are shown. Three biological replicates, n=100 cilia per sample and repetition. Data include mean ± s.d. and P values are calculated by two-tailed unpaired student t-test. h, Model depicting the role of longer distal segments on cilia stability and signalling.
Figure 3A. 3.8 Å cryoEM map of V1 in complex with subunit C, highlighting the hitherto unseen density (red) wedged between subunits B and C and the N-terminal part of peripheral stator EG2. B. Mass spectrometry analysis of V1Hchim. Protein IDs are plotted as a function of overall peptide spectrum matches (PSM) and the posterior error probability (PEP) scores. C. Left, homology model of yeast Oxr1 (residues 70-232) based on zebrafish OXR2's TLDc domain (PDB 4ACJ) (Blaise et al., 2012). Right, fit of the real space refined yeast Oxr1 homology model into the cryoEM density. D. Enlarged view of the central β sheet comprising residues Y130-K135 (β2), I141-S146 (β3), F165-K168 (β4) and A266-R271 (β8). E. Ribbon diagram of V1(C)Oxr1 featuring the binding position of Oxr1 between EG2, the C-terminal domains of the subunit AB pair adjacent to the open catalytic site, and subunit C. F. Electrostatic surface representations of V1 subunits E, A, B and C involved in Oxr1 binding.
Inactivation of endogenous Rags by starvation prevents lysosomal localization of TFEB.Figure 4.(A) ARPE-19 cells were infected with adenovirus expressing TFEB-FLAG-WT. 16 h later, cells were incubated with 250 nM Torin-1 for 1 h or starved in serum- and amino acid-free medium for 3 h. Cells were then fixed, permeabilized with 0.2% saponin, and double stained with antibodies against TFEB (used to detect recombinant TFEB) and Lamp1. (B) ARPE-19 cells were infected with adenovirus expressing TFEB-FLAG-S211A. 12 h later, cells were starved in serum- and amino acid-free medium for 4 h (starvation) or kept in complete medium (control). Cells were then fixed, permeabilized with 0.2% Triton X-100, and stained with antibodies against FLAG (used to detect TFEB-S211A). (C) HeLa cells were incubated with 250 nM Torin-1 for 1 h or starved in serum- and amino acid-free medium for 4 h with the addition of Torin-1 during the last hour of starvation. Cells were then washed, fixed, permeabilized with 0.2% Triton X-100, and double stained with antibodies against TFEB (used to detect endogenous TFEB) and Lamp1. Regions within the dotted boxes are magnified in the insets.
Ectopic expression of human S100A9 in HEK293T cells inhibits HIV infection(A, B) S100A9 expression was analysed by (A) immunoblotting or (B) flow cytometry in stably transduced HEK293T cells using anti-myc and anti-S100A9 as indicated.Figure 6S100A9 expression was analysed by (A) immunoblotting or (B) flow cytometry in stably transduced HEK293T cells using anti-myc and anti-S100A9 as indicated. Immunoblot loading controls were done with anti-actin. Flow cytometry negative and positive controls were done with non-transduced parental cells (in grey) and HEK293T cells stably expressing the red emitting fluorescent protein E2C (E2C-Far Red), respectively. Experiments shown are representative of three (n=3). HEK293T were infected with HIV-Luc for 48h (10ng of p24). Graphs represent fold RLU means ± SD of infected S100A9-expressing cells normalized to infected E2C-expressing cells from duplicates of three independent experiments (n=3). E2C- and S100A9-expressing HEK293T cells were infected with the indicated MOI of HIV-GFP for 48h and GFP+ cells were analysed by flow cytometry. Mean values ± SD from four independent experiments (n=4) were normalized to the E2C-HEK293T control and represented in graph.
Figure 3A-D (A, B) Body weight curves and (C, D) grip response - ability to grip onto an inverted wire screen for a 60-second period - in wild-type and Gaa-/- mice untreated or treated with escalating doses of AT845 (3×1013, 1×1014, and 3×1014 vg/kg). All groups had n=9 at all timepoints, except the 6 week timepoint (females) and the 12 week timepoint (males and females) for the Gaa-/- mice, which had n=8 per group. Female mice dosed with 3×1013, 1×1014, and 3×1014 vg/kg had a 22%, 11%, and 0% failure rate at week 12, respectively; in comparison, the female control WT mice had a 22% failure rate and the control Gaa-/- mice had a 38% failure. Male mice dosed with 3×1013, 1×1014, and 3×1014 vg/kg had a 56%, 33%, and 11% failure rate at week 12, respectively; in comparison, the male control WT mice had a 11% failure rate and the control Gaa-/- mice had a 100% failure. Bars represent median. Data show a dose-dependent improvement in grip response following treatment with AT845. Statistical analysis: two-way ANOVA, Dunnett's test. Asterisks (*) indicate significant differences compared with untreated Gaa-/- mice at each respective time point. *, P<0.05; **, P<0.01; ***, P<0.001; ****, P<0.0001.
Figure 4A Time course of the change in BMS scores in NB-3+/+ and NB-3−/−mice as measured with the BMS rating scale after spinal cord complete transection. *p < 0.05; two-way ANOVA followed by Bonferroni's post-test. n= 25 mice per group. B Diagram showing electrophysiological analysis of transmission across the spinal lesion site. C Electrophysiology transmission across the spinal lesion site. In each group of NB-3+/+ and NB-3−/−mice, the electrophysiological responses in the caudal spinal cord were recorded in the condition of sham-operated or 12 weeks post-injury. The latencies and amplitudes of responses were analyzed and quantified in each group. n.s., not significant, *p < 0.05, and **p < 0.01; two-way ANOVA followed by Bonferroni's post-test. n= 10 mice per group. D Diagram showing analysis of synapse reformation between regenerative axons and neurons in the injured spinal cord 0.5 mm rostral to the lesion site or 0.5 mm caudal to the lesion site. E Representative images of coronal sections showing synapse formation between the glutamatergic corticospinal terminals and spinal neurons in NB-3+/+ and NB-3−/−mice. Corticospinal axons, neurons, and glutamatergic terminals of corticospinal axons were labeled with BDA (red), Tuj1 (blue) and vGlut1/2 (green), respectively. Each group showed the data from the rostral and caudal spinal cord in the condition of sham-operated or 12 weeks post-injury. Scale bar, 20 μm.F Quantification of the number of glutamatergic corticospinal synapses per neuron in each group. n.s., not significant, **p < 0.01; two-way ANOVA followed by Fisher's LSD. n= 15 mice per group.
Figure 5(A) Cellular protein extracts were isolated from HeLa cells transfected with oligonucleotides that targeted different regions of MIA40 mRNA or with control oligonucleotides. The samples were analyzed by reducing SDS-PAGE and Western blot. (B) Cellular protein extracts were isolated from HeLa cells transfected with oligonucleotides that targeted different regions of ALR mRNA or with control oligonucleotides. The samples were analyzed by reducing SDS-PAGE and Western blot. (C) Schematic representation of MIA40 sequence containing the CPC motif in HEK293 MIA40 WT/WT and HEK293 MIA40 WT/Del53-60 cells. (D) Cellular protein extracts were isolated from HEK293 MIA40 WT/WT and WT/Del53-60 cells. The samples were analyzed by SDS-PAGE and Western blot. (E) Cellular protein extracts were isolated from HEK293 MIA40 WT/WT and WT/Del53-60 cells transfected with a plasmid that encoded MIA40 or an empty vector. The samples were subjected to reducing SDS-PAGE and Western blot. Radiolabeled [35S]COA7 (F) precursors were imported into mitochondria that were isolated from HEK293 MIA40 WT/WT and WT/Del53-60 cells. The samples were analyzed by reducing SDS-PAGE and autoradiography. The results of three biological replicates were analyzed, quantified, and normalized to control mitochondria at 45 min. The data are expressed as a mean ± SEM (n=3). IAA, iodoacetamide. Radiolabeled [ [35S]TIMM8A (G) precursors were imported into mitochondria that were isolated from HEK293 MIA40 WT/WT and WT/Del53-60 cells. The samples were analyzed by reducing SDS-PAGE and autoradiography. The results of three biological replicates were analyzed, quantified, and normalized to control mitochondria at 45 min. The data are expressed as a mean ± SEM (n=3). IAA, iodoacetamide.
Figure 4A Schematic illustration of treatment scheme. Hemizygous K18-hACE2 mice were treated intranasally with 20 µg of NM1251 (n = 15) or NM1267 (n = 12) seven hours prior to infection with 3*103 PFU SARS-CoV-2 B.1. Weight loss were monitored for 14 days. Dashed line indicates humane endpoint and symbols represent mean ± SEM ****P < 0.0001, by two-way ANOVA with Sidak's multiple comparison test in C Hemizygous K18-hACE2 mice were treated intranasally with 20 µg of NM1251 (n = 15) or NM1267 (n = 12) seven hours prior to infection with 3*103 PFU SARS-CoV-2 B.1. survival were monitored for 14 days. , ****P < 0.0001, by log-rank test D Nasal swabs were collected from six mice per group (n = 6) at the indicated time points and viral load was determined by plaque-assay. Bars represent mean ± SEM. **P < 0.01, by unpaired t test