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(F) RNA affinity chromatography using the 5'p53RNA or a dsDNA and total extracts from cells transfected with HA-tagged Ku70 wild-type (WT) or mutated (Mut6E) Ku70 [25] for 48 h, followed by western blot analysis. All the experiments were performed with HCT116 cells.
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Chaining indices of the indicated genotypes. Statistical significance was compared with control. n = 8, 23, 6 (B) Courtship indices for one target (Canton-S) male and one tester male of the indicated genotypes. Changes were determined by comparing the data with control. n = 10, 16, 21 (C) Sexual preference of single male of indicated genotypes toward a decapitated virgin female and a decapitated naïve male. Statistical significance was compared with the other gender. n = 10, 10 (D)
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A Scheme of the mathematical model. Integrated pAKT and ppERK obtained from our kinetic model were linked by linear regression analysis to measured cell-cycle indicator. Similarly, cell-cycle indicator and integrated pS6 were linked to measured proliferation by linear regression analysis and model selection.
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(E) Hek-HT-iRFP-LC3 cells were transiently transfected with siRNA targeting endogenous STK38 (or with non-targeting siRNA (siNT)). Two days after, cells were transiently transfected with indicated Flag-XPO1 mutant plasmids. 24 hours later, cells were incubated with DMEM or EBSS for 4 hours both supplemented with KPT-185 (final concentration = 1 µM) in order to inhibit endogenous XPO1 activity and with chloroquine (final concentration = 10 µM). Cells were then fixed and the number of iRFP-LC3 dots per cell was recorded only in cells positive for Flag-XPO1 mutants As expected, silencing of endogenous STK38 prevented the formation of LC3 dots upon starvation where introduction of phosphonegative XPO1 (S1055A) failed also to increase LC3 dots upon starvation. Interestingly, phosphomimetic XPO1 (S1055D) was sufficient to increase the number of LC3 dots even in complete and also STK38-depleted conditions.
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qRT-PCR data showing that CBF1 expression is induced by different light regimes. Wild-type (Col) seedlings were first grown at 22°C in D for 4 d, and then transferred to continuous W, FR, R, or B light for the indicated times ranging from 1 h to 24 h. Error bars represent SD of three technical replicates. Different letters represent significant differences by one-way ANOVA with Duncan's post hoc test (P < 0.05).
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e As in Fig. 4c but now MK-801 and saline are used for pre-treatment. The MK-801 effect was longer lasting compared to memantine and one out of four animals showed resistance to tone exposure even 4 days after MK-801 injection.
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(F) Scatter plot displaying log2 SILAC ratios of phosphorylation sites in the experiments setup SR on the x-axis and setup SR igo1∆igo2∆ on the y-axis. Phosphorylation sites that do not exhibit Cdc55-dependence are shown in grey, whereas Cdc55-dependent phosphorylation sites are highlighted in orange. We found an enrichment for Cdc55-dependent sites in the SR- and Igo1/Igo2-dependent set (Set1) (Fisher Exact test, p-value = 3.54 x 10-31, odds-ratio = 21.40).
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(E, F) qRT-PCR characterization of the AT and endothelial layers on-chip for SARS-CoV-2 entry markers (ACE2, TMPRSS2, NRP1) for cells extracted from the apical and vascular channels from uninfected LoCs (n=3 biological replicates) (E) and infected LoCs (n=4 biological replicates) at 1 day-post infection (F). In both cases expression is normalized to GAPDH levels. Data information: The bars represent the mean value, the solid line represents the median value, and the error bars represent the standard deviation. P-values are calculated using a Kruskal-Wallis one-way ANOVA test, * represents p≤0.05, ** represents p≤0.01, ** represents p≤0.001. Scale bar = 20 μm.
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Top: Schematic of the SUMO-mediated TDG-BERosome (grey oval) formation and DNA processing. Bottom: Reconstitution of active DNA demethylation on a 5caC containing synthetic DNA double-strands in presence of dynamic SUMO-conjugation/de-conjugation. Left panels lanes 3-14: BER reconstitution with XRCC1 or XRCC1-SUMO as indicated. Right panels lanes 15-21: BER reconstitution with free SUMO or XRCC1-SUMO as SUMO donor as indicated. ds59merDNA substrates: lane 1-2 CG/CG HpaII methylation sensitive restriction site, cleavable by HpaII; lane 3-6, 8-14, 15-17, 21 HpaII methylation sensitive restriction site restriction site with a caC (5caCG/CG), not cleavable by HpaII; lane 7 and 20 repaired Product (CC/CG) that is cleavable by HpaII after excision/repair of caC/CG to CG/CG by BER. Percentage of total signal of fully repaired product is indicated. ss59merDNA, single stranded DNA.
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(I) Detection of MTs and pericentrin-containing MTOCs by IF in control and RanT24N, T42A - expressing oocytes. Arrow indicates unaligned chromosome. (J) Quantification of chromosomal spread area in oocytes depicted in (I), Means ± SDs, Mann-Whitney test, oocyte numbers indicated in brackets.
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B-D. 3T3-L1 stable cells expressing vector control (EV), Flag-tagged WT or mutant WDTC1 proteins were adipogenically induced. Triglyceride levels were quantified by an enzymatic assay (C).
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(C) The effect of HOIP on Htt-Q97 aggregation is independent of NF-κB signaling. SH-SY5Y cells transiently transfected with Htt-Q97 and either control vector (CO) or WT HOIP plus the NF-κB super-repressor IκB-2S/A were analyzed by immunocytochemistry. Data are mean ± SD with n=5 from an unpaired two-tailed Student's t-test. Expression levels of WT HOIP and IκB-2S/A were analyzed by Western blotting. Data information: *p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001.
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Representative confocal micrographs (scale bars are 20 µm) of caspase 3/7 activation in DMKG (2 mM)-treated CTNSWT, CTNS‑/-, and CTNSPatient cells for 24 hrs (n = 3).
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D-E. Bar graphs showing the number of Jmjd2a peaks displaying at least 1 bp overlap with (E) a Jmjd2c peak. Jmjd2a peaks were classified as TSS-associated ('TSS') if overlapping a region of +/- 1kb of a TSS.
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GSEC structure with co-purifying helical peptide (PDB: 5FN3) is presented. PSEN1 is shown in light brown, PEN-2 in dark brown, APH1A in gold, NCT in green and the co-purifying helical peptide in red. The topology of the helical peptide (putative GSEC substrate) is indicated by the amino and carboxyl terminal groups. The interface between the N-terminal part of the helical peptide, the NCT ectodomain, and PSEN1 is magnified and NCT/PSEN1 side chains shown.
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(F) Effect of ANO8 expression on the Co-IP of the native SERCA2 and STIM1 in resting and store depleted cells. The columns are the mean±s.e.m of 3 experiments. Data information: The first number in parenthesis indicates the number of similar experiments performed and the second number is the number of cells analyzed. All results are given as mean±s.e.m of the indicated number of experiments or cells analyzed, and difference were analyzed by unpaired t test.
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A, B KidneyCoQ9 levels in Coq9+/+, Coq9Q95X and Coq9R239Xmice treated with 2,4-diHB (+2,4-diHB) compared with the non-treated littermate (vehicle). Statistical analysis was performed on +2,4-diHBCoq9+/+, Coq9Q95X and Coq9R239Xmice versus vehicle Coq9+/+, Coq9Q95X and Coq9R239Xmice, respectively (n = 3 for each group).
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D. Workflow of the strategy to map trajectories of AEC resistance using CREATE. Briefly, designed cassettes were cloned, miniprepped and transformed into strains expressing Cas9 and the lambda red machinery. The library culture was grown for 8 hours in LB media with proper antibiotics, washed with PBS, and inoculated into M9 minimal media containing the AEC selective pressure and antibiotics. An aliquot was stored for initial plasmid barcode sequencing counts. After growth, cells were harvested for deep sequencing of the plasmid barcodes, which were used to map the enrichment scores of the designed mutants
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(C) The presence of neutralizing antibodies in sera from donors #46 and #48, found negative by ELISA and positive by flow cytometry (Fig. 3C) was demonstrated as in panel B. Data represent the mean±SD of triplicates. *, p<0.05; **, p<0.005; ****, p<0.00005 (Paired two-tailed t-test comparing all serum dilutions to the pre-COVID sample).
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Virus titers of lungs were determined on Vero E6 cells (c, unpaired t-test, ***p<0.001)
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RNA FISH for uc.291 (red, Quasar 570) on A253 in scramble (SCR) and silenced (si-uc.291(1) and si-uc.291(2)) cells. Nuclei were stained with DAPI. Scale bar: 10 µM. The histogram derives from the RT-qPCR using the leftover cells after removing the slides used for the FISH staining. Data shown are mean ± s.d; n=3 biological replicates *p<0.05, Student's t-test.
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Pie chart with proteins downregulated in ECE2 KO COs sorted by relevant categories (Number in the elements = number of proteins falling into each category, with numerous proteins falling into several ones) [50-52].
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(D) Left panel: the distribution of endogenous LC3 was monitored by immunofluorescence and confocal microscopy in BI‐1 WT and KO MEFs cells at basal conditions (NT) or after exposure to EBSS for 3 h. Scale bar: left 15 μm and right 10 μm. Right panel: quantification of the number cells containing three or more LC3‐positive vesicles (N=160 cells). Mean and standard deviation are presented (N=4). Student's t‐test was used to analyse statistical significance, **P0.001 and *P0.01.
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Analysis of GAL1 loss (2-DOG resistant colonies) in tetraploid strain RBY18 after growth on PRE-SPO medium with or without antioxidants for 7 days at 37°C.
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B, C LGR5hi and LGR5lo cells were freshly isolated from 3‐month‐old LGR5‐GFPki mice 6 h after 12 Gy γ‐irradiation (IR) or from non‐irradiated (NIR) mice (n = 3 mice per group). (C) Representative Western blot analysis of cell lysates for the expression of phospho‐p53 and cleaved caspase‐3.
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A, B) Schematic depiction of ftsj1 and wdr6, and target sites of mutations introduced by the CRISPR-Cas9 system in HEK293T cells. Shaded and open boxes indicate coding regions and untranslated regions of exons, respectively. Lines indicate introns. For ftsj1, two sgRNA sequences for targeting exons 2 and 3 are noted. For wdr6, two sgRNA sequences for targeting exons 1 and 2 are noted. Sequences of both alleles of ftsj1 or wdr6 in KO1 and KO2 cell lines are aligned. Deleted nucleotides are indicated as dashed lines.
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C Time course of EN expression after partial hepatectomy (2/3 PHx) determined by qRT-PCR of total liver lysates (n = 5/time point, sham: n = 3).
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(A) SGC7901 and MGC803 cells were transfected with ATF4 shRNA or shcontrol. Western blottings and qRT-PCR were used to analyze the expression of UHMK1. Data were presented as mean ± standard deviation from three replicates. Unpaired, two-tailed statistical significance was assessed by Student's t‐test. Data information: Data represent mean ± SD. **P< 0.01, ***P< 0.001.
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(B) Internally tagged HA-Mcp3 is functional. mdm10∆ cells were transformed with the empty plasmid pYX142 () or pYX142 encoding Mdm10, Mcp3, or an internally HA-tagged Mcp3 (MDM10, MCP3, HA-MCP3). Cells were grown to an OD600 of 1.0 and spotted on YPG plates in a 1:5 dilution series. Plates were incubated for growth at 37°C for 4 days.
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HC: heavy chain. Endogenous HSF1-AMPKα interactions in immortalized Hsf1+/+ and Hsf1−/− MEFs were visualized in situ by PLA (I). Experimental details are described in Materials and Methods. Scale bars: 50 μm.
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(A) Fluorescence microscopy demonstrates that Lam6-GFP co-localizes with ERMES (Mdm34-Cherry) (yellow arrows) and also localizes to non-ERMES locations in the cell (red arrows). These additional locations co-localized with the NVJ (Nvj1-GFP) as well as with the vCLAMP (GFP-Vps39). Scale bar represents 5 μm.
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Western blot analyses of the effect of EV20/MMAF (10 nM) on the levels of HER2 and HER3 in BT474 and BTRH cells at the indicated incubation times with the ADC. The same Western was probed for HER2 or HER3 expression using two differently labelled species-specific secondary fluorescent antibodies. Quantitative representation of the Western studies shown in (E).
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Nanog>GFP and Gata6::mCherry fluorescence intensities of WT and Dax1-/- (D) cells after 3d in indicated conditions. Fraction of Gata6::mCherry positive cells in indicated genotypes and conditions after 3d Average and SD of at least two independent clones.
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Growth phenotype comparison of BRI1-mCit/bri1/ubp12i/ubp13 and BRI125KR-mCit/bri1/ubp12i/ubp13. The indicated transgenic lines were grown on DEX medium for 18 days. Scale bars: 10 mm.
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H. Percentage of released 14C-labeled fatty acids by Caco2 cells expressing shscramble* or shPkd2* (sequence 2) grown in a transwell system. n=6.
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C. Representative images of the migration phenotype in HCT116 cells with knockdown of candidate circRNAs.
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In vivo localization of mCherry-PI4Kβ1 expressed from the pPI4Kβ1 promoter in root tips of five-day-old complemented pi4kβ1 pi4kβ2 plants. The mCherry-PI4Kβ1 distribution was imaged using a Zeiss LSM880 in Airyscan Virtual Pinhole (VP) mode with the pinhole set to 2. Arrowheads, nascent cell plates decorated by mCherry-PI4Kβ1. Scale bar, 20 µm. Whole-mount immunostaining of five-day-old seedlings expressing mCherry-PI4kβ1 in the pi4kβ1 pi4kβ2 double mutant background using anti-tubulin (red) and anti-mCherry (green) antibodies, and DAPI (blue). (І), (ІІ), magnifications of regions marked in b, representing early and late cytokinetic stages. Scale bars, 20 µm.
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A,B: Wt or Mecp2 null NPCs plated on the grid of 64 MEA electrodes. Scale bars: 50 µm.
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(L) Relative anti-Y-Ae MFI in GC from immunised wildtype and Sh3gl1-/- mice. N = 11 from 2 experiments. Data show mean ± SEM.
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Strumpellin and VPS35 are in close proximity to RAB5 and RAB21. (A) Anti-biotin immunoprecipitation and immunoblot of endogenous Strumpellin and VPS35. Lysates correspond to 1% of input, n=3 independent experiments. RAB21 can be seen interacting with various WASH and retromer complexes subunits.
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Effects of DC and Beva treatment in 786-O orthoxenograft. Evaluation of percentage of necrotic area by histology, Box plots indicate median, Q1/Q3 and max/min value whiskers in Control (black), DC101 (red) and Beva (blue) treatment groups. 5-8 tumors/group were analyzed by Mann-Whitney test, and Mantel-Cox test for survival where *p < 0.05, **p < 0.01 and ***p < 0.001 and DC * p < 0.05 and Beva # p < 0.05 vs Control).
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Pie charts showing the genomic distribution of H2Bac and H3K27ac relative to whole genome, obtained from CHIP-sequencing experiments performed in WT dorsal hippocampus. The percentage of peaks in promoters, Upstream Regulatory Regions (Upstream; -1 kb to -20 kb relative to TSS), introns, exons, and intergenic regions is shown
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D. Confocal images of fibroblasts derived from a control, patient 1 and patient 2. Boxed regions are enlarged. Scale bar: 20µm. E. Bar graph showing ratios of fluorescent intensity of mitochondria delivered to lysosomes relative to that of mitochondria present in the cytosol (FL lyso/FL mito ). Statistical analysis were performed on mean ± SEM using Student's t-test (two-sided).
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C, D Top, representative sequences of cell contours during migration for 466 s of WT (C) and Arghap45−/− (D) naive T cells on 2D surface coated with ICAM-1 and CCL21. The color scale highlights each time-resolved shape. Rectangle length, 112,6 μm (WT) and 51,6 μm (Arghap45−/−). Bar, 10 µm. Botttom, three representative time-resolved shapes and their corresponding calculated contours are shown. Bar, 10 µm.
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D Heat map demonstrating enrichment of cardiac genes in GFP+ fractions (CT+, RA+) compared to GFP− fractions (CT−, RA−) at day 31.
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LDH release was measured after cold stress for 24 h. FCCP (200 µM), antimycin A (50 µM), mitoQ and decylQ (500 nM) were used as pretreatments for 30 min. Data are presented as mean ± SEM; n = 3 (J), biological replicates, ****p < 0.0001, ***p < 0.001, **p < 0.01, by one-way ANOVA followed by Dunnett's or Bonferroni's multiple comparison test. (H, I) Lipid peroxidation was measured by C11-BODIPY 581/591 after cold stress for 5 h. Representative plots are shown in (H). Quantification data (I), whose raw value were normalized by the C11-BODIPY 581/591 without cold stress of non-treatment, are presented as mean ± SEM; n = 5, biological replicates. **p < 0.01, *p < 0.05, by one-way ANOVA followed by Dunnett's multiple comparison test.
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A. H9c2 cells treated with tunicamycin (2 µg/ml, 12h) showed significantly lower survival when endogenous Nox4 was silenced (siNox4) as compared to cells treated with a scrambled siRNA (siCtl). Cell survival was restored by treatment with either guanabenz (Gbz, 5 µM) or salubrinal (Sal, 50 µM) but was unaffected by clonidine (Cld, 5 µM). n=3/group.
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Accumulation of α-synuclein oligomers analyzed by western blotting. The mutant (c13) and corrected (c22) neural stem cells (NSCs) were transduced with α-synuclein-expressing lentiviruses. At 6 days after differentiation, cells were extracted and fractionated to Triton X-100 (1%)-soluble and insoluble fractions. Shown is the representative WB image of three independent experiments. α-Synuclein oligomers are indicated with a bracket, while monomer bands are marked with an arrowhead.
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(a) Flow cytometry of T cells transduced with lentivirus expressing GFP and nontargeting control shRNA (Scr) or Lamp2a mRNA-specific shRNA (shL2A) (left), and immunoblot analysis of LAMP-2A in sorted GFP+ T cells (right). Numbers above bracketed lines (left) indicate percent GFP+ cells.
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(D) Representative DIC images of wild type (WT), ift54 and ift54 cells transformed with the deletion constructs as indicated. Arrows indicate cilia bulges. Scale bar, 5 μm.
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(C) Cell migration ability was evaluated by wound healing assay in HCT116 cells. Quantification was performed by measuring the smallest clearance distance of the wound. Data information: All results were expressed as mean ± SD of three independent experiments (n ≥ 3 per experimental condition). For figure C, statistical significance was assessed with a two-way ANOVA followed by Tukey's post hoc test. *P<0.05 and **P<0.01.
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cI- and cII-linke state 3 respiration in isolated heart, kidney and liver mitochondria (n=8/group) we added the CII substrate, succinate, to obtain CI&CII-linked respiration in presence of saturating ADP. Bar graphs represent mean±SD. Statistics: one-way ANOVA followed by Tukey"s test. Significant differences between groups (p value) are indicated on g
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i3 cortical neurons from a C9orf72-ALS patient and the paired isogenic control were differentiated for 21 days, then were immunostained for neurofilament (red), and SV2 (green) (A) Shown are representative z-stack confocal images of neurites.
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(A) Schematic representation of the experimental procedure. Catheters pre-colonized with S. aureus were allowed to form biofilms by incubation at 37ºC (for in vitro dispersion assay) or by subcutaneous implantation in mice (for ex vivo dispersion assay) before being treated in vitro with different mycoplasma strains and estimate biofilm dispersion by crystal violet staining.
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ParBF binding outside parSF on the F plasmid is compatible with a power-law decay. High resolution ChIP-seq performed on DLT3586 carrying the F plasmid (F1-10B). The ParB density, normalized to 1 at the first bp downstream the last parSF binding repeat after background subtraction, is displayed over 14-Kbp on the right side of parSF. Monte Carlo simulations and analytic formula are represented in red and dotted black lines, respectively. MC simulations were performed with a Freely-Jointed Chain of linear length L=15-Kbp and a cluster radius σ=75nm. The two other parameters, the Kuhn length a=10-bp and the total number of proteins on the F plasmid Nt=360 (related to the normalization constant of the protein concentration κ=0.41) were fitted from the ChIP-seq dat parSF inserted at the xylE locus on E. coli chromosome fro DLT207
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A) Effect of 100 nM flg22 treatment on ROS burst measured in 5 week old plants of Col-0 and mkkk7.
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E) WT PER2::LUC cells were treated with radicicol or vehicle control. Blue arrows show the time points where drug was washed off. Mean (solid lines) ± SD (dotted lines). F) Quantification of damping rate of the PER2::LUC recordings shown in E), for the duration of drug treatment. Mean±SD. One-way ANOVA with Holm-Sidak's multiple comparisons, ****p≤0.0001.
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Representative immunoblot of UCP1 protein and the loading control (Ponceau staining, PS) in iBAT samples.
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D An ATP-stabilized complex of p97 containing both UBXD1 and PLAA. Stable HEK293 cell lines were doxycycline-induced to express p97 wild-type (wt) or the ATPase-mutant E578Q (EQ) at near endogenous level. Endogenous UBXD1 was immunoprecipitated and associated proteins detected by immunoblotting with indicated antibodies. Arrowheads indicate endogenous (lower band) and induced p97 (upper band).
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C Histomorphometric analysis of trabecular bone from the femoral metaphysis. Results are shown as mean ± SEM; n=8; a: p<0.05, b: p<0.01 and c: p<0.001 by t test
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Schematic of Glycolysis and the TCA cycle. White boxes denote detected metabolites. Grey boxes denote undetected metabolites. Dashed arrows indicate presence of multiple reactions between metabolites.
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A. H&E staining of mouse organ sections. Scale bars: spleen, colon 100 μM, bone marrow - 50 μM.
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(E) Strategy for assessing the effect of increasing uSTAT5 on megakaryocytic differentiation of HPC7 cells.
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(B) LCL1.11 cells transfected with NeoR or NucNeoR expression plasmids were tested likewise. Presentation of endogenous NeoR and NucNeoR was blocked by all inhibitors, indicating that NeoR and NucNeoR presentation is dependent on autophagy and lysosomal processing.
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F Expression of Pltp mRNA in indicated tissues. Data are shown by relative Pltp mRNA levels normalized by 18s expression. n=3 for all tissues (except BAT and WAT n=5, Testis n=2). *P<0.05 Pltp expression in BAT vs other tissues (except testis and lung) by Student's t-test.
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(A) Diagram of dosing regimen for the dose-dependent effect of MOTS-c on PMO activity in mdx mice. i.v. refers to intravenous injection. PMO-M (+MOTS-c) means PMO-M supplemented with additional MOTS-c.
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(A) Vinculin was knocked down as described (Figure 5D) in MCF10A cells and cells were stained for LATS1 and TRIP6. Merged images show LATS1 (green), TRIP6 (red) and DNA (blue). Scale bar=20µm.
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Representative confocal immunofluorescence images of HA-tagged MIC19 variants with MitoTracker staining in U2OS cells. All MIC19 variants examined carried the S6W,T7P mutation. The fluorescence intensities along the dashed lines are shown as line profile graphs on the right . Scale bar = 20 µm.
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G. Z-stack gallery depicting mid-invading myoA KO parasites. Note that the nucleus is located located at the back during invasion events. Data information: Scale bar represents 5 µm. White arrow points to direction of invasion.
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WT mice (n=3-5) were treated with PBS or 12.5 mg/kg LPS with or without 10 mg/kg DEX pre-treatment. Ileum was isolated 6 hours after LPS challenge. (A) p-MLKL and MLKL protein levels and (B) RIPK3 protein levels were analyzed via western blot using ACTIN as a loading control. Expression levels were quantified using FIJI and normalized to ACTIN levels. P-values were calculated using 1-way ANOVA.
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A. Schematic of the genetic system used to analyse inducible DSB repaired by SSA. Chr. V contains ura3-52 and URA3 (at the endogenous URA3 locus) separated by ~ 4 kb of DNA containing KAN (grey box, arrow above indicates the promoter) and an HO site (triangle). Galactose-inducible expression of the HO endonuclease leads to DSB formation at the HO site. After DSB repair via SSA, the majority of cells become Kans Ura- as the 766 bp homology between URA3 and ura3-52 (grey shadows) is predominantly used.
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(A-B) Representative confocal images of liver (A) and brain (B) slices from R6/2 mice ip injected with C-NPs (left) or with g7-NPs (right) and sacrificed after 4 hrs. (C) Quantification of g7-NPs localized in the liver, striatum, and cortex of WT (n=3) and R6/2 mice (n=3). Data represent mean (µg) ± SEM.
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miR-181a/b-1-/- mice show increased mtDNA content vs. miR-181a/b-1+/+ mice as measured by q-PCR. N=4 animals/genotype.
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(C) Quantitative analysis showing distribution of GnRH neurons in the nose, OB and VFB of mutant and control embryos. Two-way ANOVA, Fisher's LSD multiple-comparison test, n=5 and 6 mouse embryos for mutants and controls, respectively, from at least 3 litters.
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C, Representative images of autonomous fluorescent signals of 4HRE-d2GFP in xenografts derived from or shUSP33- or shNT- expressing T387 GSCs transduced with 4HRE-d2GFP. Scale bar, 200 μm.
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(G, H) Representation of the phosphorylation levels at epitopes from nucleoporins (NUPs; (G)) or Retinoblastoma (RB1; (H)) upon USP7i or OA treatments from data obtained at phosphoproteomic analyses.
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Representative composite immune-fluorescent images of whole hair follicle units by confocal from non-lesional and lesional scalp psoriasis patients (D) and magnification images of specific regions from the HFs (E). CD200 (green), c-JUN (white), TSLP (red), DAPI (blue).
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(E, F) Overexpression of AAV-hTau or simultaneous downregulation of STAT1 changed the mRNA levels of GluN1, GluN2A and GluN2B detected by qRT-PCR in the hippocampal CA3. Data information: Data were presented as mean ± SD (n=4; Mann-Whitney test). *, p<0.05, **, p<0.01, ***, p<0.001 vs eGFP or hTau.
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(B) Disintegration reaction (1 hour) performed by RAG1 aa 216-1008 R848M/E649V, RAG2 aa 1-361 and full length human HMGB1 with Mg2+ or Mn2+ at the indicated temperatures. Denaturing gel displays the fluorophore-labeled DNA strand from the RAG STC before (16 nt band, lane 2) and after (37 nt) the disintegration reaction. Lane 1, fluorophore-labeled 37 nt DNA marker. Representative of 2 independent experiments.
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B. Exoproteome analysis of LukAB mutants produced by S. aureus strain Newman in a toxinless background. Samples were electrophoresed on a 12% SDS-PAGE gel and visualized by staining with Coomassie blue. A representative of two experiments is shown.
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(C) PBneutrophil production and nuclear segmentation were assessed in control and Am80-GCSF mice surviving on day 13 (section ii).
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C) WCL from WT and USP7 CRISPR KO cells were subjected to immunoblotting with different antibodies as indicated.
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B. Regulators of the balance between mesendoderm and ectoderm specification at the exit of pluripotency.
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(G) Representative stereoscopic image of WT and IκBα-deleted organoids. Scale bars in G, 50 μm.
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(A) Sequence similarities of MHR in HIV-1 (NL4-3), SIVmac239, and MuLV with the core SxIP-binding motif (bold text) within the EBH domain of EB1 from different species, adapted from (Honnappa et al., 2009). Residues identical to those in EB1 are highlighted in red, similar residues are printed in red.
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(A) The molecular species-selectivity of PS-to-PE conversion is recapitulated in wild type BY4741. WT cells overexpressing SCT1 vs. control were pulsed for 20 min with 2H3-serine prior to lipid extraction and ESI-MS/MS analysis of 2H3-PS (*PS) and 2H3-PE (*PE). Data is presented as mean of 4 biological replicates with the individual values indicated. Underlying data for this figure can be found in Dataset EV2.
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(A) Transgenic Hek293 cells constitutively expressing a separase sensor (cartoon below) and inducibly expressing Myc-separase were transiently transfected to express Flag-AsiSI-ER, Dox- and/or OHT-treated in G2-phase and analysed by (IP-)Western
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(E-G) RT-PCR analysis in circulating γδ T cells of Map3k2 or Notch2 expression (E), Nkg2d or Tnfa expression (F), and NKG2D or TNF-α protein expression (G) after siRNA-mediated knockdown of Map3k2 or Notch2. In each experiment, a control condition was also used where a nontargeting negative control siRNA was transfected. γδ were cultured with IL-7 for 4 days (n=6 independent biological samples).
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(G) Similar transfection as in (C) was performed, and the transfected cells were incubated with Bafilomycin A1 or Earle's Balanced Salt Solution (starvation) as in (D). Lysates from the cells were subjected to immunoblot analysis using anti-TDP-43, p62 and GAPDH antibodies. The relative densities are shown on the right side. The data from three independent experiments are presented as means ± S.E.M., ns, not significantly different; *, p<0.05; **, p<0.01, one-way ANOVA.
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Number of SCAP-positive ER exit sites per cell. Normalized fluorescent intensity of SCAP at each individual ER exit site. Total fluorescent intensity of SCAP at ER exit sites per cell. Number of Sec12-positive ER exit sites per cell. Data information: Data represent Mean ± SEM; NS: not significant, *: p < 0.05, **: p < 0.01, ***: p < 0.001, by 2-tailed Student's t test. In panel C-F, quantification of images from 3 independent experiments.
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] |
(C) Macrophages were pre-treated with either 1 μM GSK inh or 0.1 μM MLi-2 and then treated with 1 mM LLOMe for 30 min. Rab8A and Rab8A pT72 levels were analysed by Western blot.
|
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14,
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] |
representative HE staining image, and (P) histopathological score of colon sections from the treated WT colitis mice on day 8 (n = 8).
|
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D Quantification of emitted light intensity per plant for control and drought-treated plants.
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RT-qPCR for panel of astrocyte markers in mock and let‐7 + 125 treated Dgcr8Δ/Δ GPCs following 48 h of differentiation (n = 3).
|
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C, Spinning disk confocal microscopy on live cells reveals the elongation state of nesprins at the nuclear envelope. Dashed white lines represent the outlines of the constriction. The distance between mCh-LAC and GFP-nesprin-2 was measured at the back, at the constriction, at the front near the constriction, and at the center of the front. (N = 17 cells, over 3 biological replicates. The alignment of the two channels was corrected using at least 28 fluorescent beads.)
|
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] |
E. miRNA reverse northern blot for LincGET and Dyei. miRNAs were isolated and amplified by RT-PCR after adding double adaptors, then southern blot (reverse northern) was performed. It shows no evidence of small RNA products, indicating that LincGET and Dyei do not function as pre-miRNA. The miR1982 was used as a miRNA positive control. About 400 early 4-cell embryos were used for each experiment and three experimental replicates were performed.
|
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] |
B. Duolink proximity ligation assay between Mitofilin and CHCHD10 (upper panels) and, CHCHD10 and CHCHD6 (lower panels) in control and patient fibroblasts observed by confocal microscopy. Mitochondria were stained with Mitotracker. Duolink spots per cell were quantified for 30 randomly-selected individual cells per each studied fibroblast cell line from 2 independent experiments. Differences between the cell lines were analyzed by Student's t-test (two-sided): extremely significant (***: p=0.0001). Scale bar = 10 µm.
|
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] |
E Representative images captured by transmission electron microscopy of DC1 cells treated with vehicle or NR. White arrows: mitochondria. Yellow arrow: autophagosome-like structures with engulfed mitochondria. Enlarged image within white frame is shown. F-H Quantifications of percentage of damaged mitochondria per cell (F), mitochondrial length (G), and mitochondrial diameter (H). A minimum of 200 mitochondria counted per group.
|
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N, O Protein expression of Msps and TACC in brain tissues. (N) Larval brains of msps810/mspsP transheterozygotes at early wL3 stage were immunoblotted and probed with α-Msps and α-TACC antibody. Actin blotting was used as loading control. The dot plot on the right depicts the quantitative analysis of total TACC levels in control and msps transheterozygous larvae.
|
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] |
F. MDCK cells were cultured sEVs were isolated from the pre-cleared medium by direct immunoaffinity capture using anti-CD9 antibody.
|
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