EMBO/sd-character-level-ner huggingface dataset:
Figure 4A BR enhances the stability of BIC1 proteins. The 5-d-old 35S:BIC1-YFP transgenic plants were treated with 100 μM cycloheximide (CHX) or co-treated with 100 μM CHX and 1 μM epibrassinolide (eBL). Samples were collected at indicated time points and BIC1-YFP protein levels were analyzed by western blots using anti-GFP antibody. Actin was used to verify equal protein loadings. Intensity of the bands was measured using Adobe Photoshop CS3 Extended program. Three independent experiments were performed with similar results. B Scheme represents full length and truncated versions of BIC1. NT, amino terminal domain; CT, carboxyl terminal domain; CID, CRY interacting domain. C Transactivation analysis of BIC1 using a yeast assays. The GAL4 DNA-binding domain (BD) alone was used as the negative control. SD-W, synthetic dextrose medium lacking Trp; SD-WH, synthetic dextrose medium lacking both Trp and His. D Constructs used for BIC1 transcriptional activity assays as shown in (E). VP16 was used as a positive control. TATA, TATA box for DNA binding; LUC, firefly luciferase; REN, Renilla luciferase; NOS, Nopaline Synthase terminator. E Transient transcriptional activity analysis in Arabidopsis protoplasts illustrating the transcriptional activation activity of BIC1. The relative luciferase activities were calculated by normalizing the LUC values against REN. Error bars represent SD (n = 3). **p < 0.01, as determined by a Student's t-test. F Constructs used for BIC1 transcriptional activity assay as shown in (G). G Transient expression assays in Arabidopsis protoplasts showing that BIC1 and BZR1 synergistically activate PRE5 promoter. The ProPRE5:LUC reporter was co-transformed with the indicated effector constructs. The LUC/REN ratio represents the ProPRE5:LUC activity relative to the internal control (REN driven by the 35S promoter). Error bars represent SD (n = 3). H, I BIC1-mediated hypocotyl elongation was reduced by the addition of SRDX motif. Seedlings were grown for 5 days under long-day conditions. Images of the representative seedlings are shown in (H), and the hypocotyl lengths of the indicated genotypes were measured and are shown in (I). #1, #2, #3 represent different 35S:BIC1-SRDX-Flag transgenic lines. Data are means ± SD (n>20). **p < 0.01, as determined by Student's t-test. Scale bar, 2 mm.
Figure 1A. Root cartoon with a cell file colored to depict the MZ (green) with NMZ cells, the EZ; (yellow) with NEZ cells, and the DZ (blue), which extends more cells upwards (not shown). rMZ and rEZ are growth factors of each zone. The first and last meristematic cells, and first mature cells are encircled in black, while QC cells are in red. The Lateral root cap is not entirely depicted for simplicity. B. Upper panel. Root growth model cartoon. Numbers (i=1,2,3...) label the order at which each cell (rectangle) entered the EZ (yellow) and the DZ (blue). ti is the time at which the i-th cell entered the EZ. Because of the irregularity in mitotic events at the meristem. ti - ti-1 is set to be stochastic. The simulations start with a single cell in the EZ. The EZ grows until it reaches the stationary regime, when it will be composed of nearly the same number of cells over time. All results in figures correspond to the stationary regime. Lower panel. Simulation of cell length exponential growth for three different cells (colors) of three simulated root files. Results herein correspond to the Sizer model. C. Cell length profile as a function of the cell number position from the QC for three wild-type seedlings (symbols, top) at day 8 post germination and three simulated root files for the Sizer model (symbols, bottom). Notice the use of a logarithmic scale in cell lengths, which visualizes the cell lengthlog10 rEZ
TgAtg3 conditional knock out mutant.Figure 7A. Schematic representation of the strategy. Recombineered cosmid bearing chloramphenicol acetyl transferase (CAT) gene for selection of resistant parasites, flanked by homologous regions from TgAtg3, was used to replace endogenous genomic locus in the cell line expressing a regulatable extra copy of TgAtg3 (imyc-sTgAtg3). Mutants obtained after double homologous recombination events were selected with chloramphenicol. PCR amplifications and Southern blot strategy used for subsequent characterisation of the mutant are indicated in red and green, respectively. B. PCR verification of the removal of TgAtg3 in the mutant (imyc-sTgAtg3-ΔAtg3, A) compared to the parental cell line (imyc-sTgAtg3, B) C. Southern blot verification of the TgAtg3 gene replacement. ∼4 µg of genomic DNA from mutant (A) and parental (B) tachyzoites were digested by HindIII/AvrII, transferred on a nylon membrane and probed with 32P-labeled 5′ and 3′ DNA probes located as schematised in A.
Figure 4A Maximum intensity projections of confocal z-stacks of 4-day-old animals expressing the indicated transgenes or harboring the indicated temperature-sensitive (ts) mutations. Muscle cells were stained with anti-myosin (green) and Alexa Flour 647-phalloidin (purple). Scale bar: 5 µm. Myosin misfolding reflects the exposure of the ts mutant phenotype of myosin(ts) [unc-54(e1301)] at the permissive temperate of 15°C by co-expression of the hsp-110 hairpins. B Quantification of motility as a measure for toxicity. Displayed is the mean ± SEM (N = 4) of body bends/30s of animals expressing the indicated transgenes or harboring the indicated mutations. Statistical analysis was done using one-way ANOVA with Dunnett's multiple comparison test. *** p ≤ 0.001. Co-expression of the hsp-110 hairpins exposed the ts mutant phenotype at the permissive temperate of 15°C, resulting in a significant increase in movement defects. All strains contain the sid-1(pk3321) allele. C Quantification of motility as a measure for transgene toxicity. Displayed is the mean ± SEM (N = 4) amount of body bends/30s of animals expressing the indicated transgenes or harboring the indicated mutations. Statistical analysis was done using two-way ANOVA with Tukey's multiple comparison test. * p ≤ 0.05. Expression of the hsp-110 hairpins caused movement defects with increasing age. D Maximum intensity projections of confocal z-stacks of 12-day-old animals expressing the indicated transgenes. Scale bar: 10 µm. FLUCSM forms foci with increasing age, which is markedly accelerated upon hsp-110 KD. E Quantification of animals with FLUCSM::GFP foci at indicated days of life. Data are displayed as mean ± SEM (in %) (N = 3). Statistical analysis was done using two-way ANOVA with Dunnett's multiple comparison test. n.s. = not significant, ** p ≤ 0.01, *** p ≤ 0.001. F Quantification of motility as a measure for transgene toxicity. Displayed is the mean ± SEM (N = 3) amount of body bends/30s of animals expressing the indicated transgenes or harboring the indicated mutations. Statistical analysis was done using two-way ANOVA with Dunnett's multiple comparison test. n.s. = not significant, *** p ≤ 0.001. Expression of the hsp-110 hairpins caused movement defects with increasing age. The beneficial effects of HSP-110 KD on amyloid toxicity is masked in older animals. G Quantification of motility as a measure for transgene toxicity. Displayed is the mean ± SEM (N = 3) amount of body bends/30s of animals expressing the indicated transgenes or harboring the indicated mutations. Statistical analysis was done using two-way ANOVA with Dunnett's multiple comparison test. n.s. = not significant, , ** p ≤ 0.01, *** p ≤ 0.001. Expression of the hsp-110 hairpins caused movement defects with increasing age. The beneficial effects of HSP-110 KD on amyloid toxicity is masked in older animals. Data information: All strains harbor the sid-1(pk3321) allele.
Characterization of FBXO7-/- mice.Figure 1A. Lysates of indicated tissues isolated from P30 rat were immunoblotted with the FBXO7 and γ-tubulin antibodies. The latter served as loading control. B. Schematic of the genetically modified PARK15 gene locus located on chromosome 22. C. Brain lysates from P5 FBXO7+/+, FBXO7+/- or FBXO7-/- mice were immunoblotted with the FBXO7, βgal, γ-tubulin or pan 14-3-3 antibodies. The latter two served as loading controls. D. LacZstaining of P18 FBXO7-/- sagittal brain section. Ctx = cortex, Hpc = hippocampus, Cb = cerebellum, Olf = olfactory bulb, CC = corpus callosum, Str = striatum, Th = thalamus. Scale bar = 1 mm. E. and F. Body weight of P5 or P18 FBXO7+/+, FBXO7+/- or FBXO7-/- mice. n=17, 46, 16 (E) and n=16, 25, 11 (F), respectively (ANOVA, ***p<0.001, mean + s.e.m.). G. Images of representative P18 FBXO7+/+, FBXO7+/- or FBXO7-/- mice. H. Kaplan-Meier survival curve of male and female FBXO7-/- mice. n=14 and 16, respectively. I. and J. P18 FBXO7+/+, FBXO7+/- or FBXO7-/- littermates were tested in the open field (I) and on the rotarod (J). n=20, 23, 16, respectively (ANOVA, ***p<0.001, mean + s.e.m.). K. Average weight of 6 month and 12 month-old FBXO7+/+ and FBXO7+/- littermates. n=15 and 18, respectively (ANOVA, mean + s.e.m.). L. 6 month and 12 month-old FBXO7+/+ and FBXO7+/- littermates were examined on the rotarod. n=15 and 18, respectively (ANOVA, mean + s.e.m.).
Figure 4(A) Schematic representation of the experimental setup for investigating the influence of overactive exopheresis on F0 and F1 worm generation. (B) Exopher production does not improve muscle functionality. n = 11 - 38; N = 3. (C, D) Animals with overactive exopheresis have reduced exploratory behaviour presented as a reduction in straight line distance travelled (C) and by representative worms traces on the plate (D). n = 11-34; N = 3. (E) RNAi knockdown of the egg yolk precursor protein VIT-1 increases the number of muscular exophers. n = 20; N = 2. (F) Embryos from hermaphrodite mothers that produce a high number of exophers contain more egg yolk precursor protein VIT-2. Representative images of embryos with endogenous VIT-2::GFP levels from mothers with different exopheresis activity are shown above the graph. n = 15 and 14, N = 5 and 6. Scale bar is 10 µm. (G) RNAi knockdown of RME-2 yolk receptor abolish exopher production. n = 110-123; N = 3. (H) Muscle-produced VIT-2 is released from muscles via exophers. The image shows the midbody of worms expressing the proteasome subunit RPN-5 tagged with RFP in BWM and VIT-2::GFP endogenous expression. Arrows: white - exopher, blue - muscle cell. Dashed lines mark embryos present in the uterus. Scale bar is 10 µm. (I) Offspring of worms with overactive exopheresis develop faster. Y.A. - young adult stage, L4 - last larval stage. n = 317-372; N = 2.
Characterisation of proviral integrations in corrected stem cells.Figure 5Visualisation of COL7A1 sequence by FISH analysis in corrected stem cells at passage XII (clone 6). Five specific signals (red) were detected on five different chromosomes. As expected, a COL7A1 probe (red) hybridised to the two endogenous COL7A1 alleles located on the chromosomes 3 as identified by a specific centromeric probe (Cen 3) and to three other chromosomes identified as chromosomes 2, 11 and 22 by means of specific centromeric probes (Cen 2, Cen 11 and Cen 22, respectively). The latter corresponded to proviruses.Determination of proviral integration sites in corrected stem cell. Genomic DNA from clone 6 was submitted to whole-genome sequencing together with PCR and subsequent Sanger sequencing to identify the integration sites to the base-pair level. Mate pairs that span from the viral sequence to a human chromosome were extracted and used to estimate integration regions. The reads were mapped to the hg19 reference sequence. Three integration sites were uncovered: one in chromosome 2, one in chromosome 11 and one in chromosome 22, and targeted genes were identified. Analysis of the level of expression of targeted genes in corrected stem cells by quantitative RT-PCR. DARS was not significantly changed in clone 6 compared to the cells before transduction. Primers used annealed downstream of the proviral integration site. Error bars represent the standard deviation of three replicates.Identification of sequence abnormalities in the two alleles of the DARS-targeted gene in clone 6 by NGS. Reads were mapped to the hg19 reference sequence. Small insertion-deletions and SNP calling were performed and compared to GWAS databases. The mapping highlighted two indels and one SNP in the DARS genomic sequence. The indels were not associated with disease and the SNP was synonymous (CCU-CCG both codons correspond to proline).
Localization of GFP-Atg8 during UPR Induction.Figure 7Localization of GFP-Atg8 during UPR InductionSome of the DTT-treated cells shown in Figure 4B expressing GFP-Atg8 and Sec61-cherry (as an ER marker) were visualized using fluorescence microscopy. GFP-Atg8 localizes in close proximity to the ERAs detected by the ER marker.
Figure 4(A) qPCR analysis of proinflammatory cytokines Il-6, Tnfα, Il-1β and antiviral cytokine Ifnβ in WT cultured cortical and hippocampal neurons. Data were analyzed by unpaired t-test. Il-6 and Tnfα N=8; Ifnβ and Il-1β, N=6. (B, C) Poly(I:C) treatment inhibits dendrite outgrowth of Il-6-/- neurons (B) and Tnfα-/- neurons (C). Mean values SEM of representatives of three independent experiments are shown. The numbers of analyzed neurons in the representative experiments are indicated in each column. Data were analyzed by unpaired t-test. (D) Left, schematic of experiment using conditioned medium. Upper right, conditioned medium applied to WT neurons. Lower right, conditioned medium applied to Tlr3-/- neurons. Dendrite morphology was analyzed one day after adding conditioned medium. Mean values SEM of representatives of three independent experiments are shown. The numbers of analyzed neurons in the representative experiments are indicated in each column. Data were analyzed by unpaired t-test. * P < 0.05, *** P < 0.0001.
Fig.Figure 3A) Gene set enrichment analysis using Fisher's exact test of GWAS genes from Marioni et al. (Marioni et al, 2018) at different p-value cut-offs among the WGCNA-derived modules in APPtg or TAUtg mice Colors represent -log10 (Benjamini-Yuketieli adjusted p-value) for the enrichment. Numbers on the x-axis: black = p-value cut-off; grey = size of GWAS gene set. B) Log2 fold change (LFC) in TAUtg (x-axis) and APPtg mice (y-axis) after differential expression analysis, assessing the effects of age*genotype interaction. Color code for dots/numbers: grey = genes in hippocampus (n=15824); black = genes in APPtg-Blue (n=4236); green = genes significantly differentially expressed in APPtg mice (n=493); yellow = significantly differentially expressed genes in TAUtg mice (n=9); red = significantly differentially expressed genes in both (n=9). C) Z-score distribution per experimental group for all genes within the APPtg-Blue module. Boxplots: center line, median; box limits, 25th-75th quartiles; whiskers, 1.5x interquartile range. Empirical p-values are Bonferroni adjusted (***pbonf<0.001) and indicate significant shift in z-score distribution D) Gene Ontology (GO) enrichment for genes within the APPtg-Blue module. The x-axis depicts -log10(FDR-adjusted p-value) multiplied by the enrichment score (ES), where the red line represents an -log10(0.049)*(ES=1). E) The "top 18" GWAS genes are prioritized by finding the intersection of genes within the APPtg-Blue module (n=4236), AD GWAS genes with p<0.001 in Marioni et al. (Marioni et al, 2018) (n=314) and significantly differentially expressed genes in the age*genotype comparison of APPtg mice (n=798), using the SuperExactTest (Wang et al, 2015).
Figure 7A. Graph representing the percentage of adult C. elegans animals with 6 normal dopaminergic neurons. Heterozygous expression with hVPS41WT (strains UA386 or UA387) or compound heterozygous expression of hVPS41 variants (strain UA388) does not yield significant changes in neurodegeneration compared to the Pdat-1::GFP (strain BY250) control at either day 7 or day 10 post-hatching. n=30 Adult worms for each of 3 independent experiments for GFP (total of 90 worms) and n=90 for each independent transgenic strain (30 worms/trial x 3 independent transgenic lines = 270 worms), for each of 3 independent experiments. B. Representative images of the neuroprotection assay described in (A), where worms express GFP specifically in the anterior 6 DA neurons. Intact DA neurons are indicated with arrowheads. hVPS41 variant backgrounds are expressed as indicated. Scale bar 20µm. C. Graph representing the percentage of animals with 6 normal dopaminergic neurons in the anterior region of Pdat-1::GFP; Pdat-1:: α-syn (strain UA44) animals with heterozygous expression of hVPS41 variants. Heterozygous expression of hVPS41WT with either variant (strains UA389 or UA390) significantly rescues neurons from α-synuclein-induced degeneration at day 7 whereas compound heterozygous expression (strain UA391) fails to rescue neurodegeneration. At day 10, none of the heterozygous backgrounds significantly rescues α-synuclein-induced neurodegeneration. n=30 Adult worms for each of 3 independent experiments for the α-synuclein (α-syn) strain (total of 90 worms) and n=90 for each independent transgenic strain (30 worms/trial x 3 independent transgenic lines = 270 worms) for each of 3 independent experiments. D. Representative images of DA neurons from C. elegans expressing Pdat-1::GFP; Pdat-1:: α-syn, with or without hVPS41 variants, as described in (C). Intact DA neurons are indicated with arrowheads and missing neurons are indicated with arrows. Scale bar 20µm.
| The EV-vsiRNAs confer antiviral activity upon antiviral RNAi activation VsiRNAs are induced in virus-infected cells by dicing the viral double-stranded RNA replicative intermediates and loaded into RISC for subsequent cleavage of viral RNA genomes.Figure 6VsiRNAs are induced in virus-infected cells by dicing the viral double-stranded RNA replicative intermediates and loaded into RISC for subsequent cleavage of viral RNA genomes. Part of the vsiRNAs enter the EVs together with miRNAs, ncRNAs, proteins, and lipids. EVs in MVBs are secreted to the extracellular system. When attached to recipient cells, vsiRNAs in EVs are released into the cytoplasm to restrict the virus replication.
Fig.Figure 5WT or LRRK2 KO RAW264.7 macrophages, RAW264.7 macrophages pre-treated with 1 μM GSK2578215A (GSK inh) and WT or Rab8A KO RAW264.7 macrophages were treated with 1 mM LLOMe for 30 min. LC3B positive vesicle numbers were analysed by immunofluorescence and high content imaging. Scale bar = 20 μm. Right panels show quantification of number of positive vesicles per cell. Data show the mean ± SEM of three to four biological replicates. ns=non-significant, *p≤0.05, **p≤0.01 by One-way ANOVA followed by Sidak's multiple comparisons test. WT or LRRK2 KO RAW264.7 macrophages, RAW264.7 macrophages pre-treated with 1 μM GSK2578215A (GSK inh) and WT or Rab8A KO RAW264.7 macrophages were treated with 1 mM LLOMe for 30 min. ubiquitin K63 positive vesicle numbers were analysed by immunofluorescence and high content imaging. Scale bar = 20 μm. Right panels show quantification of number of positive vesicles per cell. Data show the mean ± SEM of three to four biological replicates. ns=non-significant, *p≤0.05, **p≤0.01 by One-way ANOVA followed by Sidak's multiple comparisons test. (C) Representative images of LC3B/LAMP-1 double positive endolysosomes in control and LLOMe treated macrophages. Scale bar = 10 μm. (D) Analysis of the % of LC3B/LAMP-1 double positive endolysosomes in LLOMe treated macrophages. Data shows mean ± SEM of three independent biological experiments. *p≤0.05, ***p≤0.001 by Student's t-test.
Identification and characterization of COI genesLife cycle of Tetrahymena.Figure 1A Life cycle of Tetrahymena. A single cell of Tetrahymena thermophila contains two different types of nuclei: a macronucleus (MAC) and a micronucleus (MIC). When sufficient nutrients are available, Tetrahymena grows by binary fission, and the MAC and MIC are divided independently (vegetative growth, green). After prolonged starvation (blue), two cells of complementary mating types fuse to begin the sexual reproduction process (conjugation, pink). Their MICs undergo meiosis (a), and one of the meiotic products survives and divides mitotically, giving rise to two pronuclei: one stationary and one migratory (b). The migratory gametic nucleus crosses the conjugation bridge (c) and fuses with the stationary nucleus to produce the zygotic nucleus (d). The zygotic nucleus divides twice (e). Two of the products differentiate into the new MACs, whereas the other two remain as MICs (f). The parental MAC is degraded, and the pair is dissolved (g). The exconjugants resume vegetative growth when nutrients are available (h). The approximate time at which each event occurs in our culture condition is indicated (hpm: hours post‐mixing). B The expression of the predicted Tetrahymena genes showing more than 19 expressed sequence tags (ESTs) from conjugating cells but not vegetative cells was analyzed by RT-PCR using total RNAs from exponentially growing vegetative cells ("E"); 16‐h starved cells ("S"); or conjugating cells at 2, 4, 6, 8, 10, and 12 hpm. Twenty‐two genes that were exclusively expressed during conjugation are marked with a yellow background. Eleven genes that were analyzed by gene knockout (KO) are indicated in the "KO" column. The phenotypes are described in the "KO phenotype" column. The predicted TTHERM_00526270 contains two genes, which were analyzed separately in this study. C Exconjugants of wild‐type (WT) and KO strains for the indicated COI genes at 36-48 hpm were used to detect Tlr1 IES elements by fluorescence in situ hybridization (FISH). The percentages of total exconjugants that showed severe (++), mild (+), and no (−) DNA elimination defects are given (n > 100). Representative pictures of exconjugants showing each phenotype are shown above. The Tlr1FISH signal is in red, and the DNA was stained with DAPI (blue). The MICs (i) and the new MACs (na) are marked.
Figure 1A. Quantitative analysis of developmental expression of Lrig1 mRNA in rat hippocampus by real-time RT-PCR. The results are shown as mean SEM of n=3 independent assays. The levels of Lrig1 mRNA were normalized using the expression of the housekeeping gene Tbp (TATA binding protein). The insert shows the expression of Lrig1 in embryonic E17.5 rat hippocampus examined by RT-PCR. Control sample without reverse transcriptase (-RT) is also shown. B. Localization of Lrig1 (red) in coronal sections from P15 ratbrain by immunofluorescence. Anti-Lrig1ECD antibodies label dentate gyrus cells, CA1-CA3 hippocampalneurons and pyramidal corticalneurons (*). Scale bar, 400 m. C-E. Confocal image of cortical (C) and hippocampal (D-E) pyramidal neurons stained with rabbit anti-Lrig1ECD antibody. Arrows indicate Lrig1 staining in proximal segments of apical dendrites of CA1-CA3hippocampal and pyramidal cortical neurons (layer V). Scale bars, 20 m. F. Immunofluorescence staining of Lrig1 (red) with anti-Lrig1ECD antibody and the neuronal marker ßIII tubulin (green) in dissociated hippocampal cells cultured for 7 DIV. Yellow indicates neuronal expression of Lrig1. Scale bar, 15 m. G. Immunofluorescence staining of Lrig1 (red) with anti-Lrig1ECD antibody and the astrocytic marker S100 (green) in dissociated hippocampal cells cultured for 7 DIV. Scale bar, 20 m. H. Localization of Lrig1 (red) with anti-Lrig1ECD antibody and the somatodendritic marker MAP-2 (blue) by immunocytochemistry in dissociated rat hippocampalneurons after 12 DIV. Scale bar, 20 m.