EMBO/sd-character-level-ner huggingface dataset:
Figure 6(a) IHC analysis for the microglia marker Iba1 in Emx1-Cre/Trim28-KO animals and controls. Scale bars: Low magnification 75 μm, high magnification 10 μm.(d) Western blot revealed an increased expression of CD68 in the Emx1-Cre/Trim28-KO animals (n=5 per group), unpaired t-test, **p=0.0036, error bars mean ± SEM. The star indicates an unspecific band. (e) An increased expression of the ERV derived protein IAP-Gag was detected in the cortex of Emx1-Cre/Trim28-KO animals (n=5 per group), unpaired t-test, ***p=0.0008, error bars mean ± SEM. (f) Immunohistochemistry for IAP-gag visualized the presence of ERV proteins in the cortex of Emx1-Cre/Trim28-KO animals. The distribution of IAP-gag was heterogenous among Trim28-KO neurons, where it was either accumulated in a low (#) or high (°) number of aggregate-like structures in the cytoplasm, or as weak homogenous staining throughout the cytoplasm (#) or not present at all (*). Scale bars: low magnification 75μm, high magnification 5 μm.
a) Western blots of lysates from the indicated cell lines.figf4(a) Western blots of lysates from the indicated cell lines. (b) Pearson correlation analysis of p27 and LKB1 protein levels quantified by reverse phase protein array assays in 74 randomly selected primary breast tumors (MDACC). (c) Western blots of protein lysates from Hela cells transfected with either pCDNA3 or wild-type LKB1 cultured with or without glucose for 24 h. (d-f) Western blots of protein extracts from MEFs treated with AICAR (d and e), (d-f) Western blots of protein extracts from MEFs treated with AICAR (d and e) (d-f) Western blots of protein extractsfrom 3T3L1 cells cultured with or without glucose for 30 min (f). (g) Western blot analysis of 3T3L1 cells 72 h post-transfection RNAi with or without glucose starvation for 1 h before analysis. Densitometric quantification of western blots is shown. Uncropped images of the blots are shown in the Supplementary Information, Fig. S5.
Figure 6Based on the structures of apo-PfFNT and PfFNT-MMV007839, we propose that the PfFNT membrane protein is capable of picking up lactate ions from the parasite's cytosol, converting them to lactic acids and then exporting these acids into the extracellular space via a sequential displacement mechanism. We believe that residues lining the central channel are highly flexible and can quickly rearrange sequentially to contract and relax the channel for propelling the export of substrate across the membrane. This schematic diagram indicates that PfFNT shuttles lactate ions from the parasite's cytoplasm to the midway of the central channel. These ions will be converted to lactic acids, via proton transfer from H230, and then shuttled across the second half of the channel to the parasite's extracellular surface. Residue K35 contributes to form a transient binding site to bind lactate ion nearby to the cytoplasmic surface of the parasite. As soon as the second lactate ion arrives from the parasite's cytoplasm, the first lactate will be displaced and continue to propagate to the midway of the channel, where H230 is responsible for donating a proton to convert this lactate ion into lactic acid and open the constriction for further propagation. Residue K177 participates in forming a transient binding site, which binds lactic acid nearby the extracellular surface. A second ion enters this site will cause displacement and extrusion of lactic acid out of the parasite.
Crystal structure of Munc18-1 containing KKEE mutation.Figure 7A Ribbon diagrams of the structure of rat Munc18-1 containing the K332E/K333E mutation, two orientations rotating 90° with respect to each other were shown. Two Munc18-1 K332E/K333E molecules in an asymmetric unit pack against each other through helix-helix interaction in domain 3a. B,C Structure alignment of domain 3a with various SM proteins. Crystal structure of domain 3a from rat Munc18-1 K332E/K333E was aligned and superposed with rat Munc18-1 WT in complex with closed Syx1 (pdb entry: 3C98) (B), and with human Munc18-2 (pdb entry:4CCA), nSec1 from squid (pdb entry:1FVH), and rat Munc18-1 WT in complex with mouse Syx4 N-peptide (pdb entry: 3PUJ) (C). D The flexibility of SM proteins in domain 3a and domain 1. Structures of rat Munc18-1 WT in complex with closed Syx1, human Munc18-2, nSec1 from squid, and rat Munc18-1 WT in complex with mouse Syx4 N-peptide were aligned and superposed with rat Munc18-1 K332E/K333E. Enlarged views of domain 3a and domain 1 are shown in dashed boxes.
Fig.Figure 2(A) Cytosolic ATP synthesis in the indicated VSMC types, measured by incorporation of phosphate-32 (32Pi) into ADP. [32Pi]-ATP was separated from 32Pi using the molybdate method, as explained in the Methods section. (B) Intracellular lactate concentration (C) external acidification in the indicated VSMC types. (D) Calcium accumulation in mitochondria after 24 h of incubation in MEM media containing 10 µCi/mL calcium-45 (45Ca2+) as a radiotracer. (E) Magnesium concentration in isolated mitochondria. (F) The boxed scheme describes the mitochondrial calcium overload hypothesis. Lactic acidosis forces the Na+/H+ exchanger (NHX) to import Na+, resulting in cytosolic Na+ overload. Subsequently, the Na+/Ca2+ exchanger (NCX) is forced into reverse mode to dispose of excess Na+, resulting in cytosolic calcium overload. This Ca2+ is then taken up by mitochondria, resulting in mitochondrial calcium overload. 2-DG (2-deoxyglucose; 50 mmol/L) blocks glycolysis through competitive hexokinase inhibition, whereas oligomycin (10 µmol/L) inhibits mitochondrial ATP synthase. G-6-P: glucose-6-phosphate.
Munc13 promotes Syb2 binding to the Munc18-1/Syx1 complex.Figure 5A Schematic depiction of Syb2 binding to Munc18-1/Syx1 complex in the presence of MUN domain. The extended instead of bent conformation of domain 3a within the Munc18-1/Syx1 complex is compatible with Syb2 binding. B Effects of the QKAA, KKEE, and L348R mutations on binding of Syb2 to Munc18-1/Syx1 complex in the presence and absence of MUN domain detected by GST pull-down assay. Bands of Munc18-1 are indicated with red asterisks.
Figure 6Extraction of cortices revealed similar human tau (Tau13) in TBS-extracts (not significant) but significantly more human tau in TritonX-100 (TX-100) extracts of rTg4510-Mapt0/0 compared to rTg4510 mice. Mean±SEM, n =3 mice/group, two-tailed Student's T test, ns non significant. Native gel electrophoresis of cortical TBS-extracts showed small differences in HMW (oligomeric) human tau between rTg4510 and rTg4510-Mapt0/0 brains. Western blot lanes were averaged across ~2/3 of the width (black rectangular and arrow in Tau13 blot). The mean±SEM (n=3 mice/group) of these averages was plotted as longitudinal lane profiles. Differences in HMW tau are indicated by red and black arrows. TBS-brain extracts were applied to a HEK293 cell tau aggregation seeding assay(Holmes et al, 2014, Sanders et al, 2014), in which TauRDP301S-CFP and TauRDP301S-YFP are co-expressed intracellularly. The formation of intracellular fluorescent TauRDP301S aggregates leads to Foerster resonance energy transfer (FRET) activity between co-aggregated CFP and YFP-tags and correlates with the tau aggregation seeding activity of the applied brain extracts. After 24 hours, cells treated with extract (0.5 µg and 1.0 µg total protein per 96 well) from 9-month-old rTg4510 had significantly more intracellular YFP-positive (white arrows) aggregates compared to rTg4510-Mapt0/0 extracts; FRET activity of TauRDP301S aggregates appeared similar for both rTg4510 and rTg4510-Mapt0/0. WT and Mapt0/0 extracts never showed seeding activity. Addition of lipofectamine corrected for differences in cellular uptake of tau and led to similar differences in seeding activity between rTg4510 and rTg4510-Mapt0/0 mice. Two-tailed Student's T-test, mean±SEM ns, not significant, Insets 100 µm, Scale bars 50 µm.
Figure 6(A) Dissected mouse aorta demonstrating lipid-rich plaques in brachiocephalic artery after 4 months of high fat diet compared to the normal diet in Apoe-/- mice. (b) (d) Cross-section of brachiocephalic artery from (a) (c) stained with Oil Red O. L: lumen. Scale bar: 4 mm. 3 mice per group. (B) Histological analysis of mouse normal artery or atherosclerotic plaque in brachiocephalic artery with anti-SM -actin, anti-Notch3, and anti-SM-MHC antibodies. Nuclei were counterstained with DAPI (blue). L: lumen. Scale bar: 62 m. 3 mice per group. (C-F) Analysis of brachiocephalic artery of Apoe-/- mice maintained for 4 months on either normal or high fat diet using anti-CD31 (green), anti-p-FGFR1 (red), anti-FGFR1 (red), anti-p-Smad2 (red), and anti-p-Smad3 (red) antibodies. Nuclei counterstained with DAPI (blue). Scale bar: 62 m. L: lumen. M: Media. 6 mice per group. (G-J) Quantification of the number of medial smooth muscle cells expressing p-FGFR1, FGFR1, p-Smad2, and p-Smad3 (***p<0.001 compared to ND, NS: not significant compared to ND; unpaired two-tailed Student's t test.). ND: Normal diet. HFD: High fat diet.A full table of p-values for this figure is shown in Appendix Table S1.
Figure 1A. Left: Telomerase activity measured using ddTRAP assay. The horizontal pink lines represent the threshold used in these samples for background correction. Each dot on the ddPCR output represents a unique droplet that is either positive or negative for fluorescent signal. Right: Quantification of TRAP activity after normalization to SW39/TEL+ cells (n=3). Data information: Data are presented as mean ± SD. Statistical analyses were performed with Student's two-tailed unpaired t-test with Welch's correction for normality of distribution unless specified otherwise. PTM, post-translational modification. B. hTR and hTERT expression, normalized to ACTB mRNA and to SW39/TEL+. Data information: Data are presented as mean ± SD. Statistical analyses were performed with Student's two-tailed unpaired t-test with Welch's correction for normality of distribution unless specified otherwise. PTM, post-translational modification. C. RNA Pol II binding at hTR promoter in SI14/TEL+, SI24/ALT+ and 8G12/ALT+ cell lines, normalized to H3 ChIP. Control loci with, respectively, high or low transcriptional activity: c-fos promoter and subtelomeric 4qHox locus (n=3). Statistical analyses were performed using one-way ANOVA with post-hoc Tukey's test for multiple comparisons. Data information: Data are presented as mean ± SD. Statistical analyses were performed with Student's two-tailed unpaired t-test with Welch's correction for normality of distribution unless specified otherwise. PTM, post-translational modification. D. H3K9me3 and H3K27me3 (left) or H3K4me3 and H3Ac (right) densities at hTR promoter in SI14/TEL+ and SI24/ALT+, normalized to H3 (n=3). Control loci as in C. Data information: Data are presented as mean ± SD. Statistical analyses were performed with Student's two-tailed unpaired t-test with Welch's correction for normality of distribution unless specified otherwise. PTM, post-translational modification. E. NFYB binding at hTR promoter in SI14/TEL+, SI24/ALT+, SW39/TEL+ and IMRB/ALT+ cells normalized to H3. Control loci with, respectively, high and low NFYB binding: hnRNPA1 promoter and subtelomeric 4qHox (n=3). Data information: Data are presented as mean ± SD. Statistical analyses were performed with Student's two-tailed unpaired t-test with Welch's correction for normality of distribution unless specified otherwise. PTM, post-translational modification.
Figure 7Rotavirus RNAs have a 5′ cap but not a 3′ poly(A) tail. Its NSP3 protein (instead of PABP) binds to the 3′ end of the viral mRNA and interacts with eIF-4G directly to maintain the closed-loop RNA for the initiation of viral RNA translation and to block host mRNA circularization (Piron et al, 1998)." Picornavirus RNA has an internal ribosome entry site (IRES) in the 5′ untranslated region and a 3′ poly(A) tail. Viral protease (2A or leader) digests eIF-4G. The truncated eIF-4G shows higher efficiency in the IRES-driven translation than the cap-dependent translation (Ohlmann et al, 1995; Ali et al, 2001). Coronavirus RNAs have a 5′ cap and a 3′ poly(A) tail. In our model, SUD associates with the 40S/80S ribosome and enhances the PABP:Paip1 interaction to stimulate the host translation machinery. Meanwhile, coronaviral Nsp1 specifically cleaves host mRNAs (green) but not viral RNAs (Kamitani et al, 2006; Huang et al, 2011). Also, viral Nsp1 blocks host mRNA binding to the 40S ribosome. As a result, SARS-CoV could increase the viral RNA translation but inhibit host mRNA translation.
A) Exogenous expressed LAMP-2A or Hsc70 co-localized with exogenous Htt-552-18Q and Htt-552-100Q in HeLa cells.Figure 2(A) Exogenous expressed LAMP-2A or Hsc70 co-localized with exogenous Htt-552-18Q and Htt-552-100Q in HeLa cells. Cells were transfected with Htt-552 for 48 h and fixed with methanol, blocked, and processed for double immunofluorescence with antibodies against LAMP-2A (green), Hsc70 (green) and Htt-552 (red). Merged images of both channels are shown at the bottom. N: nucleus. Thin arrows point to Htt-552 immunoreactivity. Thick arrows point to lysosomes. The scale bar = 10 µm. (B) Co-immunoprecipitation of exogenous expressed LAMP-2A with exogenous Htt-552. Upper panel: PC12 cell were transfected with PCDNA4-LAMP-2A for 72 h. Cell lysates were subjected to immunoprecipitation with anti-Htt antibody or no primary antibody and Western blot analysis was performed with anti-LAMP-2A antibody. Lower panel: PC12 cell were transfected with Htt-552 for 48 h. Cell lysates were subjected to immunoprecipitation with anti-LAMP-2A antibody or no primary antibody and Western blot analysis was performed with anti-Htt antibody. (C) Co-immunoprecipitation of exogenous expressed Hsc70 with Htt-552 from PC12 cells. Upper panel: PC12 cell were transfected with PCDNA4-Hsc70 for 72 h. Cell lysates were subjected to immunoprecipitation with anti-Hsc70 antibody or no primary antibody and Western blot analysis was performed with anti-Htt antibody. Lower panels:PC12 cell were transfected with Htt-552 for 48 h. Cell lysates were subjected to immunoprecipitation with anti-Htt antibody or no primary antibody and Western blot analysis was performed with anti-Hsc70 antibody. Ab: Antibody.
Figure 6Total GSH in cytosol and mitochondria of cerebrum of Coq9+/+ and Coq9R239X mice(A). Cytosolic GPx and GRd activities in cerebrum of Coq9+/+ and Coq9R239X mice (B). Levels of GPx4 (C) and GRd (D) protein in cerebral homogenate of Coq9+/+ and Coq9R239X mice. Levels of L-Glutamate (L-Glu), N-Acetyl-Glutamate (NacGlu), L-Tryptophan (L-Trp), 5HIAA, N-Acetyl-Tryptophan (NALT), L-Tyrosine (L-Tyr) in cerebrum of Coq9+/+ and Coq9R239X mice (E). Biosynthetic pathway of GSH, serotonin and catecholamine (F).
hnRNPA2-D290V causes C.Figure 6A) Top: schematic depicting hrpa-1 gene and a loss-of-function deletion allele (tm781), referred to as hrpa-1(∆). Bottom: to create a chimeric protein (HRPA-1HsLC), the third coding exon (blue) was replaced with a codon-optimized sequence encoding the majority of the human LC. B) After 22 hours of paraquat induced oxidative stress, hrpa-1(∆) animals showed increased degeneration of tail glutamatergic (phasmid) neurons, based on defective dye uptake. Defective dye uptake indicates either neuron process degeneration or death. This defect is rescued by introduction of hrpa-1HsLCWT, but not hrpa-1HsLCD290V or hrpa-1HsLCempty. N= 6-12 animals/genotype/trial, significance from two tailed t-test. C) The C. elegans ortholog of TDP-43, tdp-1, contains 7 exons. A new deletion allele, tgx58, deletes the entire coding region of tdp-1. A second deletion allele, ok803, removes part of the 4th exon and the remaining three exons. D) Loss of C. elegans TDP-43 ortholog, tdp-1, rescues glutamatergic tail/phasmid neurodegeneration caused by hrpa-1HsLCD290V after 22 hours paraquat induced oxidative stress, but not hrpa-1(∆). N=4-12 animals/genotype/trial, significance from two tailed t-test; tdp-1∆=tdp-1(tgx58). E) Loss of C. elegans TDP-43 ortholog, tdp-1, rescues glutamatergic tail/phasmid neurodegeneration caused by hrpa-1HsLCD290V after 22 hours paraquat induced oxidative stress, but not hrpa-1(∆). N=4-12 animals/genotype/trial, significance from two tailed t-test; tdp-1∆=tdp-1(ok803). F) A constitutively activated Y531F Fyn (Fyn*) construct was expressed in the glutamatergic sensory tail (phasmid) neurons using the osm-10 promoter. An empty construct removing the beginning of Fyn and causing a frame shift mutation was constructed similarly. G) Expression of Fyn* in sensory glutamatergic neurons (osm-10 promoter, unc-54 3'UTR) reduces glutamatergic tail/phasmid neuron degeneration in hrpa-1HsLCD290V animals after 22 hours paraquat induced oxidative stress. N=9-12 animals/genotype/trial, significance from two tailed t-test. H) Expression of an empty Fyn control (Fynempty) in sensory glutamatergic neurons (osm-10 promoter, unc-54 3'UTR) does not alter glutamatergic phasmid neuron degeneration in hrpa-1HsLCD290V animals after 22 hours paraquat induced oxidative stress. N=9-12 animals/genotype/trial.
Influence of rolling on single-receptor gating kinetics.Figure 6Representative recordings of patches containing one single wild-type (WT) or GluN1-E698C/GluN2B-L795C receptor. The bottom trace is an expanded view. C, closed channel; O, open channel Single channel properties (equilibrium channel open probability, mean open time, mean closed time) of the WT and super-active GluN1-E698C/GluN2B-L795C receptors Closed (left) and open (right) time histograms for WT (upper panels) or super-active GluN1-E698C/GluN2B-L795C (lower panels) receptors. Closed time histograms were best fit with 5 exponentials, whereas open time histograms were best fit with 2 exponentials (see Online Methods and Appendix Fig S8). Smooth lines are associated exponential fits Kinetic schemes and equilibrium constants for WT (upper panel) or super-active GluN1-E698C/GluN2B-L795C (lower panels) receptors. Records were analyzed at equilibrium. Ligand-binding steps are shown in gray. Note that for simplicity, long-lived desensitized steps are not shown. *P<0.05 relative to WT GluN1/GluN2B (two-tailed Student"s t-test, unpa Proposed mechanism for the conformational switch in full-length tetrameric GluN1/GluN2B receptor: inter-dimer rolling in the ABD layer is coupled to entry of the NTD layer in its active state. The positive allosteric modulator spermine, which binds the interface between GluN1 and GluN2B NTD lower lobes, enhances receptor activity by stabilizing the NTD compact form and therefore rolling. Conversely, the negative allosteric modulators zinc and ifenprodil inhibit receptor activity by stabilizing an expanded form of the NTDs (lower lobes further apart) thus preventing rolling. The rolling motion facilitates pore opening by acting on the ABD-TMD connecting linkers
is a target site of calcineurin.Figure 3CSF extract was supplemented with mRNA encoding Flag-Cdc20 that was either wild-type or mutated to serine at Thr64, Thr68 and Thr79 (3TS). After expression, calcium was added, samples were taken at the indicated time points and resolved by either conventional or Phos-tagTM SDS-PAGE. Samples were immunoblotted for the Flag-tag, Cdc27 and cyclin B2. CSF extract was supplemented with Myc-XErp1 wt IVT and treated with DMSO or CsA. Calcium was added, samples were taken at the indicated time points, resolved by either conventional or Phos-tagTM SDS-PAGE and immunoblotted for Cdc20, Cdc27 and cyclin B2. A control sample was treated with λ-phosphatase. A pseudocoloured representation of the Cdc20 and the Cdc27 immunoblots is shown in Fig. EV4A. The cyclin B2 membrane was stripped and reprobed for α-tubulin. CSF extract was supplemented with Flag-Cdc20 wt IVT coupled to α-Flag beads. Calcium was added and Flag-Cdc20 was isolated by α-Flag immunoprecipitation at the indicated time points. Input and pellet (IP: Flag) samples were immunoblotted for the Flag-tag, pThr68 Cdc20 and p150(Glued). The pThr68 Cdc20 membrane was stripped and reprobed for cyclin B2. Asterisk in the cyclin B2 immunoblot indicates the IgG heavy chain. Asterisks in other immunoblots indicate unspecific bands. CSF extract was supplemented with Flag-Cdc20 wt IVT. Flag-Cdc20 was re-isolated by α-Flag immunoprecipitation, supplemented with calcium and treated with recombinant hs_CaN and calmodulin as indicated. Samples were taken at the indicated time points and immunoblotted for the Flag-tag and pThr68 Cdc20. The pThr68 Cdc20 membrane was stripped and reprobed for the catalytic calcineurin subunit CnA. Asterisk indicates the IgG heavy chain.
IL-2 signaling supports PMA/I-inducible DHS formation A,B.Figure 4DNase-Seq tag density plots showing all peaks detected in replicate 1 of TB IL-2 and TB+ IL-2 (A, 38645) and in TB+ IL-2 and TB+ IL-2nil (B, 36067), ordered by increasing fold change in tag count as indicated. The position of the 1015 IL-2-dependent inducible DHSs is shown alongside. Average DHS signal at the 1015 IL-2-dependent inducible DHSs in TB IL-2, TB+ IL-2, TB+ IL-2nil and naïve T cells stimulated with PMA/I (TN+). De novo identification of DNA consensus motifs enriched at the 1015 IL-2 iDHSs as determined by HOMER. Bar graph showing the number of IL-2 iDHSs found within 25 kb of an IL2 pDHS. Examples of IL-2 iDHSs which are proximal to IL-2 pDHSs at the Csf2/Il2 and Il10 loci. IL-2 iDHSs are highlighted in red and IL-2 pDHSs in blue.
Figure 6Western blot results showing the levels of CPEB1 in WT and Cnot6l-/- oocytes at the indicated time points after meiotic resumption. Total proteins from 50 (A oocytes are loaded in each lane. DDB1 (A is blotted as a loading control. Experiments were performed three times with reproducible results; a representative result is shown Western blot results showing the levels of CPEB1 in WT and Cnot6l-/- oocytes at the indicated time points after meiotic resumption. Total proteins fro 100 (B) oocytes are loaded in each lane α-tubulin (B) is blotted as a loading control. Experiments were performed three times with reproducible results; a representative result is shown Immunofluorescenc 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 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 E: Relative mRNA copy numbers of polysome-bound transcripts in WT and Cnot6l-/- oocytes at indicated stages. Error bars indicate values of the replicates F: Scatter plot comparing polysome-bound transcripts between WT and Cnot6l-/- oocytes at GV, MI and MII stages, respectively. Transcripts decreased or increased more than five folds in Cnot6l-/- oocyte samples were high-lighted with blue or red, respectively G: Pie chart in left depicts total transcripts significantly degraded during GV-MI transitions in WT oocytes. Pie chart in right shows the polysome-association of transcripts that were deemed to be degraded during GV-MI transitions in WT oocytes, but were stabilized after Cnot6l knockout H: Pie chart in left depicts total transcripts significantly degraded during GV-MII transitions in WT oocytes. Pie chart in right shows the polysome-association of transcripts that were deemed to be degraded during GV-MII transitions in WT oocytes, but were stabilized after Cnot6l knockout I: Quantitative RT-PCR results showing the relative levels of indicated transcripts in association with polysomes in WT and Cnot6l-/- oocytes at GV, MI and MII stages. Error bars, standard deviations (n = 3 biologica repeats)