EMBO/sd-character-level-ner huggingface dataset:
a,b) Cells expressing either WT or D620N GFP-VPS35 were treated with siRNA to silence expression of endogenous VPS35, fixed and labelled for GFP and FKBP15 (a) or FAM21 (b).f3(a,b) Cells expressing either WT or D620N GFP-VPS35 were treated with siRNA to silence expression of endogenous VPS35, fixed and labelled for GFP and FKBP15 (a) or FAM21 (b). Cells were imaged by fluorescence microscopy. Scale bar, 20 μm. (c) Cells stably expressing WT or D620N GFP-tagged VPS35 were treated with siRNA to abolish expression of endogenous VPS35 and subsequently fixed and labelled for GFP and either VPS26, FKBP15 or FAM21. The cells were imaged using Cellomics automated fluorescence microscopy. Approximately 250 cells per well in four wells were analysed for each cell line. The data for FKBP15 and FAM21 spot intensity were normalized to GFP-VPS35 and VPS26 signals. P0.0002 for both FKBP15 and FAM21 spot intensity in D620N compared with WT. Error bars indicate s.d. (d) Cells stably expressing either WT GFP-VPS35 or the D620N mutant were treated with siRNA to silence the expression of endogenous VPS35. Cells were permeabilized by flash freezing followed by rapid thawing and centrifuged to separate supernatant (S) and membrane pellet (P) fractions. (e) The graph shows the percentage of membrane-associated FKBP15 and strumpellin and is the mean of three experiments. The error bars indicate s.e.m. A representative blot of three experiments is shown, indicating the efficacy of the knockdown of endogenous VPS35 and also further showing that membrane proteins such as the CIMPR are detected only in the pellet fraction.
Figure 3A UMAP plots focusing on DN3a, DN3b γδ+ and DN4 γδ+ cells from Dataset 1. Clustering was performed using a 1.5 resolution. The UMAP plot is colored according to clusters and the main characteristics of the defined clusters specified in the right margin. B Dot plot showing the expression level of the top 6 DEG markers characterizing each of the 12 clusters. Dot color represents the scaled average expression of the specified gene across the 12 clusters, and dot size indicates the percentage of cells expressing the specified gene. C UMAP plots colored according to the expression level of the specified genes.
Figure 1A. mATG8s specific peptides are developed analogous to ubiquitin chain specific sensors (van Wijk et al, 2012). B. Peptides are displayed on major coat protein p8 or minor coat protein p3, pre-adsorbed on non-target mATG8 (purple box) and selected on target mATG8 (blue box). Specific peptides are identified by ELISA and subsequently sequenced (green box). Efficacy and specificity of the selected sensors is determined by pull-downs, intracellular co-translocation, peptide binding-arrays and cellular assays. Sensors are further improved by rational optimization of avidity and binding affinity. C. Phage displaying specific peptides for mATG8 are identified by phage ELISA on immobilized GST-mATG8s and negative control proteins BSA and GST. Binding signal is shown as absorbance at 450 nm in arbitrary units [AU]. D. Pull-down of mTurquoise-peptide fusions from cell lysates on GST-mATG8s beads followed by immuno-blotting with α-GFP. As loading control, the protein staining with amido black is shown.
Figure 3CHME3 treated with negative control, EB1 or CLIP170 siRNAs were either lysed and analyzed by WB Molecular weight markers (in kDa) are shown to the right of WBs in A. CHME3 treated with negative control, EB1 or CLIP170 siRNAs were infected with GFP-Vpr/S15-tomato-labeled VSV-G pseudotyped HIV-1 (B). Scatterplot of maximum p24 intensity of fused viral particles normalized to the control mean. (C-D) Amounts of pelleted p24 CA normalized to the input CA at 3 h.p.i. with HIV-1-VSV-luc in 293A (C) or CHME3 (D) treated with control, EB1 or CLIP170 siRNAs. Mean +/- SEM from three independent experiments is shown. Statistical significance was determined by one-way ANOVA. Error bars = standard error of the mean (* = p-value ≤0.05, ** = p-value ≤0.01, *** = p-value ≤0.001).
Figure 3Images and comparisons of absolute and relative lacteal lengths in jejunum between vehicle- or ABX-treated mice at P7, P14, and P28. Each dot indicates mean value of 5-10 villi in a mouse (n = 6 mice/group). Scale bars, 100 μm. Images and comparisons of number of Prox1+ LECs within initial 100 μm portion in CD31+/LYVE-1+ lacteals of jejunum from vehicle- or ABX-treated mice. Dotted lines outline villi. Each dot indicates mean value of 5-10 villi in a mouse (n = 6 mice/group). Scale bars, 100 μm. Images and comparisons of proportions of each VE-cadherin LEC junctions in CD31+/LYVE-1+ lacteals in jejunum between vehicle- or ABX-treated mice at P7, P14, and P28. Button-like (red arrowheads) and zipper-like (open arrowheads) junctions are indicated in each magnified box in right (n = 6 mice/group, 5-10 villi/mouse). Scale bars, 25 μm.
Figure 1A Schematic representation of CD45 exon 5 splicing and CTCF binding in B cell lines and primary T cells, as previously characterized. B Bisulfite sequencing of CD45 exon 5 DNA in cell types that either include or exclude exon 5 from CD45 mRNA, as indicated. Closed versus open circles indicate the locations of methylated or unmethylated CpGs, respectively. A partial sequencing chromatogram from naïve peripheral T cells demonstrates both bisulfite conversion efficiency and the persistence of methylated cytosines within the CTCF binding site. C Southern blot of genomic DNA double-digested with BsaHI (methyl-sensitive) and BglI (methyl-insensitive) hybridized to a radiolabeled 5' probe. The absence of BsaHI digestion in BL-E5(+) cells confirmed CTCF binding site methylation. D Methylated DNA immunoprecipitation (MedIP) with antibodies to 5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC) in BL41 B cells sorted and maintained based on distinct exon 5 inclusion levels. qPCR with primers against the CD45 gene body, as indicated. E 5hmC MedIP as in D, but with peripheral human CD4+ T cells sorted on the basis of cell-surface expression of exon 5 containing CD45 isoforms.
Figure 3Split-ubiquitin yeast two-hybrid assay revealing the UBP13 and BRI1 interactions. The indicated constructs were cotransformed into yeast cells. BRI1-Cub with Nub, Nub-CIPK8, and Nub-UBP13 with an empty vector were used as negative controls. Transformants were streaked onto solidified medium supplemented with X-gal. Blue patch indicates positive interaction. Cub, the C-terminal half of ubiquitin; Nub, the N-terminal half of ubiquitin. UBP13 interaction with BRI1 in plants. BRI1-Myc and FLAG-UBP13 were coexpressed in N. benthamiana leaves and coimmunoprecipitation was done on solubilized microsomal proteins with α-Myc antibody beads. The association of BRI1-UBP13 was detected by western blot with an α-FLAG antibody after α-Myc IP (Top). Deubiquitinating activity of UBP13 against K48- and K63-linked ubiquitination. Diubiquitins linked through K48 or K63 (K48 Di-Ub or K63 Di-Ub) were incubated alone or with GST-UBP13 or GST-UBP13C207S for 18 h. Deubiquitination of the diubiquitins was analyzed by western blot with the α-ubiquitin (α-Ub, P4D1) antibody. The presence of GST-UBP13/UBP13C207S recombinant proteins was confirmed by the α-GST antibody. UBP13 deubiquitination of K63-ubiquitinated BRI1 in vitro. Polyubiquitinated MBP-BRI1CD was generated by incubation with GST-PUB13 (Ub reaction) and then with GST-UBP13 or GST-UBP13C207S (DUB reaction-MBP pull-down, input). After in vitro deubiquitination, MBP-BRI1CD was purified by an MBP pull-down to analyze the BRI1CD ubiquitination (DUB reaction-MBP pull-down, SE). The ubiquitinated proteins were detected by western blot with an α-ubiquitin antibody (FK2). Ubiquitin chain specificity was detected with an α-K63Ub antibody (Apu3). The presence of recombinant proteins was confirmed by the α-GST and α-MBP antibodies. Western blot analysis of BRI1 ubiquitination on plants expressing pBRI1:BRI1-mCitrine complementing the bri1 mutant in either the UBP12/UBP13 (BRI1-mCit/bri1) or ubp12i/ubp13 (BRI1-mCit/bri1/ubp12i/ubp13) background. BRI1-mCitrine proteins were isolated from 18-day-old plants grown on DEX medium and then immunoprecipitated with α-GFP antibody beads from solubilized microsomal proteins. Ubiquitinated BRI1 and basal BRI1 proteins were detected by the α-ubiquitin (P4D1) and α-GFP antibodies, respectively. Quantification of BRI1 ubiquitination profiles from (E) (n = 3 biological replicates). The ubiquitinated BRI1 fraction was normalized to the total immunoprecipitated BRI1 detected by the α-GFP antibody.
Figure 1(A) Schematic representation of BRCA1 and BARD1. Highlighted are RING domains (orange), PALB2-interacting coiled-coil region (CC; red), BRCA1 C-terminal domains (BRCT; green) and ankyrin repeats (ANK; pink). (B) Immunoblotting of whole cell extracts obtained from parental and RAP80-/- clones in the RPE-1 hTERT p53-/- Cas9, RPE-1 hTERT p53-/- BRCA1-/- Cas9, U2OS, U2OS-2-6-3 and U2OS Flp-In/TREx cell lines with RAP80 antibodies. Tubulin is used as loading control. Representative of at least two independent immunoblots. See Figure EV1 for further details on the gene editing strategy used to knockout RAP80. (C, D) The indicated parental and RAP80-/- cell lines were processed for immunofluorescence 1 h post-irradiation (10 Gy) and stained with antibodies against ABRAXAS1 and γH2AX. Shown in (C) is the percentage of cells with > 5 ABRAXAS1 foci that colocalize with γH2AX. A minimum of 100 cells per replicate were analyzed and the bars represent the mean ± S.D. (n=3 biological replicates). Representative micrographs of RPE1-hTERT p53-/- Cas9 wild type (WT) and RAP90-/- cells are shown in (D). (E, The indicated cell lines were processed for immunofluorescence 1 h post-irradiation (10 Gy) and stained with antibodies against BRCA1 and γH2AX. Shown in (E) is the percentage of cells with > 5 BRCA1 foci that colocalize with γH2AX. A minimum of 100 cells per replicate were analyzed and the bars represent the mean ± S.D. (n=3 biological replicates for all conditions except n=5 for U2OS WT, n=2 for U2OS 2-6-3 cells, n=7, 5, 4 for U2OS Flp-In/T-REx cells). F) The indicated cell lines were processed for immunofluorescence 1 h post-irradiation (10 Gy) and stained with antibodies against BRCA1 and γH2AX. Representative microgaphs of RPE-1 hTERT p53-/- Cas9 wild type (WT) and RAP80-/- are shown in (F). (G) RPE-1 hTERT p53-/- Cas9 parental (WT) and RAP80-/- cells were untreated (t=0 h) or irradiated with a 10 Gy dose. Samples were collected at the indicated time points, processed for immunofluorescence and stained with antibodies against BRCA1 and γH2AX. Shown is the percentage of cells with > 5 BRCA1 foci. A minimum of 100 cells per replicate were analyzed and the datapoints represent the mean ± S.D. (n=3 biological replicates). All scale bars are 5 μm.
Figure 7 tRNA modifications orchestrate parasite development at a translational level in a collective model whereby: (1) High levels of Am and s2U at ring stage stabilize the "hypomodified" tRNA molecules or serve as a precursor for the subsequent modifications; (2) a synchronous increase in 22 of the 28 modifications serves to signify tRNA maturation. This parallels up-regulation of translational and metabolic activities through the ring-to-trophozoite transition depicted by the graphs showing peaks of synthesis of DNA, RNA and protein (adapted from (de Rojas & Wasserman, 1985)); (3) Increase in the levels of particular wobble modifications enhance the recognition to specific codon(s) out of other synonymous codons. This results in selective translation of codon-biased transcripts which manifests in the codon usages of the late trophozoite and schizont proteomes
Figure 3(A, B) Endogenous CAMK2B redistribution to ER membrane structures labeled by mCherry-FAM134B and BAP31 upon Tg treatment. Scale bars, 10 μm. The scale bars in the magnification boxes are 2 μm. The colocalization was analyzed by Pearson's correlation coefficient (PCC) in (B). For Ctrl, 16 cells were counted(n=16); for Tg treatment, 18 cells were counted (n=18). ***P<0.001, one-way ANOVA, error bars indicate SEM. (C) In vitro FAM134B S151 phosphorylation by CAMK2B was detected by Western blot. Recombinant proteins for FAM134BWT and FAM134BS151A were purified from E. coil were incubated with purified CAMK2B by IP in kinase buffer. FAM134B phosphorylation was analyzed by a specific antibody recognizing phos-Serine151 of human FAM134B. (D) FAM134B S151 phosphorylation in cells treated with CAMK2 activator (100nM EB1089 for 1h) or/and inhibitor (10μM KN93 for 2h). 293T or SKN-SH (a cell line derived from neuroblastoma) cells were treated with drugs as indicated, whole cell lysates were analyzed by phospho-FAM134B (S151) antibody. (E) FAM134B S151 phosphorylation in CAMK2B knockdown 293T cells. (F) In vitro reconstitution of CAMK2B-FAM134B-mediated membrane fragmentation using liposome assay. Purified recombinant FAM134B and CAMK2B were preincubated in kinase buffer with ADP or ATP at 30°C for 10min, the resultant protein mixtures were transferred to chamber coated with liposomes. Liposome fragmentation was monitored by live imaging. Scale bars, 10 μm. ***P<0.001, one-way ANOVA, error bars indicate SEM (n=3). (G, H) Measurement of the ER-phagy activity. U2OS cells transiently expressing mCherry-EGFP-FAM134B (WT) were treated with compounds as indicated. Lysosomal mCherry-positive but GFP-negative puncta were quantified for each cell in (H). For vehicle control, 53 cells were counted (n=53), for Ionomycin, n=56; for EB1089, n=54; for KN-93, n=58. Scale bars, 10 μm. The scale bars in the magnification boxes are 2 μm. ***P<0.001, one-way ANOVA, error bars indicate SEM. (I) FAM134B WT or knockout (KO) U2OS cells were treated with CAMK2 activator Ionomycin (Iono), EB1089 or inhibitor KN93 for 24 hours, Western-blot was performed to analyze the proteins as indicated. (J) CAMK2B WT or knockdown (KD) U2OS cells were treated with 1 μM Thapsigargin (Tg) to induce ER stress for different time as indicated. Cells were collected and analyzed for proteins as indicated by Western blot. (K) GFP-cleavage assay. CAMK2B WT or knockdown (KD) U2OS cells were co-transfected with FAM134B-Flag and GFP-SEC61B, 24 hours post transfection, cells were treated with drugs as indicated and analyzed for GFP-cleavage by Western blot. Note: the levels of Flag-FAM134B, CAMK2B, GFP-SEC61B and Tubulin were analyzed by Western blot using different membranes.
Figure 5(A and B) In vivo limiting dilution assay results of MGC-803 cells treated with phosphate-buffered saline, BBI608, sEVs from control cells (Con sEVs), and sEVs from LSD1 knockout (KO) cells (KO sEVs) as indicated for 28 days in BALB/c-nu mice. Representative images of tumors excised from the mice (A) and the frequency of tumor formation (B) are shown (the solid line represents the sphere formation ability curve, while the dashed line represents the 95% confidence interval. The circles and triangles represent data from different groups). (C) Tumor volume of each group subjected to in vivo limiting dilution assay with 5×106 cells with indicated treatment (n = 6 biological replicates; mean ± standard error of mean(SEM), *P = 0.0441 and ***P = 0.0002; two-tailed unpaired Student's t-test). (D) Tumor weight of each group subjected to in vivo limiting dilution assay (n = 6 biological replicates; mean ± SEM; ***P < 0.0001, *P = 0.0285, *P = 0.0162, *P = 0.0498, and *P = 0.0049; two-tailed unpaired Student's t-test). (E) Expression levels of LSD1, CD44, SOX2, and OCT4 in tumor tissues subjected to in vivo limiting dilution assay performed with 5×106 cells. (F) Immunofluorescence image and quantification of the expression of SOX2 in tumor tissues subjected to in vivo limiting dilution assay. Scale bar = 50 µm (n = 3 biological replicates; mean ± SEM; **P = 0.0024, **P = 0.0088, and *P = 0.0342; two-tailed unpaired Student's t-test). (G) Representative images of second-generation tumors (left) and the tumor formation rate (right) in each group. (H) Second-generation tumor weight in each group (n = 6 biological replicates; mean ± SEM; *P = 0.0091; two-tailed unpaired Student's t-test). (I and J) Expression levels of LSD1 in sEVs isolated from the plasma. CD9 and CD63 were used as markers of sEVs. CD9 was used as a loading control for sEV lysis (n = 3 biological replicates; mean ± SEM; *P = 0.0256 (patient 1), **P = 0.0068 (patient 2), **P = 0.0013 (patient 3), **P = 0.0031 (patient 4), ***P = 0.0009 (patient 5), **P = 0.0012 (patient 6), ***P = 0.0007 (patient 7), ***P = 0.0010 (patient 8), ***P = 0.0001 (patient 9), and *P = 0.0461 (patient 10); two-tailed unpaired Student's t-test). (I and J) Expression levels of LSD1 in sEVs isolated from the plasma. CD9 and CD63 were used as markers of sEVs. CD9 was used as a loading control for sEV lysis (n = 3 biological replicates; mean ± SEM; *P = 0.0256 (patient 1), **P = 0.0068 (patient 2), **P = 0.0013 (patient 3), **P = 0.0031 (patient 4), ***P = 0.0009 (patient 5), **P = 0.0012 (patient 6), ***P = 0.0007 (patient 7), ***P = 0.0010 (patient 8), ***P = 0.0001 (patient 9), and *P = 0.0461 (patient 10); two-tailed unpaired Student's t-test). (K) Sphere formation assay results of MGC-803 cells treated with sEVs as indicated for 7 days. Scale bar = 100 µm (n = 3 biological replicates; mean ± SEM; **P = 0.0019 and **P = 0.0050; two-tailed unpaired Student's t-test).
Figure 1A, Cell viability of NS, GBM, and astrocyte cell lines measured 72 h after irradiation (fold vs. non-irradiated cells, ctrl). *: one-way ANOVA, P=0.0001. B, Cell viability of BT308NS and their pseudo-differentiated progeny (BT308 diff) measured 24 h after irradiation (fold vs. non-irradiated cells, ctrl). *: t-test, P=0.0006. C, Flow cytometry analysis of phosphorylated histone H2AX (H2AX) at the indicated time points after IR (5 Gy, fold vs. non-irradiated cells, ctrl). *: one-way ANOVA, P=0.03. D, Neutral comet assay performed 24 h after IR (5 Gy). Ctrl: non-irradiated cells. Comet tail's length is proportional to DSB extent. E, Western blot showing phosphorylation of ATM (pATM) and Chk2 (pChk2), and accumulation of RAD51 after IR (5 Gy). Total ATM protein is also shown. Vinculin was used as loading control. F, qPCR of RAD51 expression measured in NS, BT308 diff, and astrocytes at the indicated time-points after IR (5 Gy, fold vs. non-irradiated cells, ctrl). Data are represented as mean ± SEM.
Pgs1 depletion rescues mitochondrial fragmentation by inhibiting mitochondrial fission.Figure 4Equal amounts of protein extracted from total (T), cytosolic flow-through (C) and mitochondrial eluate (M) from MEFs of the indicated genotypes stably expressing MitoTag (pMX-OMP25-GFP-HA) obtained following mitochondrial immunocapture were separated by SDS-PAGE, immunoblotted with indicated antibody and quantified by densitometry. Data represent mean ± SD of three independent experiments, One-way ANOVA. Representative confocal images of MEFs of the indicated genotypes showing subcellular Drp1 distribution. Mitochondria (Tom40, green), Drp1 labelled with anti-Drp1 antibody (red) and nuclei (NucBlue, blue). Scale bar=10μm. MiD49/51/Mff KO MEFs lack all 3 Drp1 receptors (MiD49, MiD51, and Mff). Bar graph representation of Drp1 localized to mitochondria (green) vs cytosol (blue). Data represent mean ± SD of three independent experiments, (884-3116 cells per cell line), unpaired t-test; *p < 0.05. Representative confocal images of live cell imaging of MEFs of the indicated genotypes subjected fragmentation with 5μM carbonyl cyanide m-chlorophenyl hydrazine (CCCP) for the indicated time points. Images were captured every hour for 18 hours. Scale bar=10μm. Supervised ML mitochondrial morphology quantification using WT MEFs treated with 5μM CCCP for 18h (fragmented), untreated (normal), or treated with 10μM CHX for 9h (hypertubular) training sets. Data represent mean ± SD of three independent experiments, (131-426 cells per cell line), One-way ANOVA; *p < 0.05, ** p < 0.01, ****p < 0.0001, ns; not significant. FRAP fusion assay in MEFs of the indicated genotype (see Supplemental Movies 1-3). Scale bar=10μm. Quantification of mitoYFP signal intensity measured at 200 ms intervals in the photobleached area (green box) for the indicated time (seconds), represented as relative fold recovery post-bleach. Data represent mean ± SEM of two independent experiments (n=18-52 cells per genotype), One-way ANOVA. Representative confocal images of live cell imaging of MEFs of the indicated genotypes subjected hyperfusion (SiMH) with 10μM cycloheximide (CHX) for the indicated time points. Images were captured every hour for 9 hours. Mitochondrial morphology quantification of using WT MEFs treated with 5μM CCCP for 18h (fragmented), untreated (normal), or treated with 10μM CHX for 9h (hypertubular) training sets. Data represent mean ± SD of four independent experiments, (155-745 cells per cell line), One-way ANOVA.
Figure 5A Western blot showing the expression of macroH2A proteins in control HepG2 cells, knock-down for macroH2A1 and macroH2A2 (DKD) and DKD with ectopic expression of the indicated YFP-tagged constructs. H3 is used as a loading control. B H3K9me3 immunostaining in the indicated HepG2 cell lines with nuclear counterstaining in DAPI. The indicated areas are zoomed-in for a better observation of the H3K9me3 signal profile. All images are maximum intensity Z-projections of confocal stacks. Quantification of the nuclear area The box limits correspond to 25th and 75th percentiles, the bold line indicates median and cross indicates the average values. (*, p<0.05 using unpaired two-tailed Student's T-test; n.s., not significant). Quantification of H3K9me3 foci per nucleus, The box limits correspond to 25th and 75th percentiles, the bold line indicates median and cross indicates the average values. (*, p<0.05 using unpaired two-tailed Student's T-test; n.s., not significant).
Figure 7Double immunolabeling of SYCP3 (red) and either γH2AX (pseudocolored in blue in Pds5AB cKO spread spermatocytes at the indicated stages. Double immunolabeling of SYCP3 (red) and RAD51 (green in Pds5AB cKO spread spermatocytes at the indicated stages. Data information: Sex bivalents (XY) are indicated. Scale bar, 10 μm. I Scatter dot-plot graph of the quantification of RAD51 foci in in 15 zygotene nuclei randomly selected from three different individuals of each genotype. Bars indicate mean and standard error and statistical significance was assessed with an unpaired two-tailed T-test function (****, p < 0.0001). J-L Double immunolabeling of MLH1 (pseudocolored in white) and SYCP3 (red) in Pds5AB cKO spread spermatocytes. Data information: Sex bivalents (XY) are indicated. Scale bar, 10 μm.
Prevention of DCM phenotype development in adult Ttn Ser14450fsX4 knock-in mice by injection of vivo-morpholino-modified antisense oligonucleotides.Figure 6Experimental design. Echocardiographic analysis of adult knock-in mice. Statistical difference was tested using the two-sided Student's t-test (WT: n = 6, HET: n = 6; HET + vPMO-mScrAON: n = 10 and HET + vPMO-mTtnAON: n = 14; *P = 0.013, HET versus HET + vPMO-mTtnAON and *P = 0.02, HET + vPMO-mScrAON versus HET + vPMO-mTtnAON for LVEDD differences; **P = 0.006, HET versus HET + vPMO-mTtnAON and *P = 0.02, HET + vPMO-mScrAON versus HET + vPMO-mTtnAON for LV-EF differences; *P = 0.03, HET versus HET + vPMO-mTtnAON and *P = 0.04, HET + vPMO-mScrAON versus HET + vPMO-mTtnAON for IVSd differences; **P = 0.001, HET versus HET + vPMO-mTtnAON and *P = 0.03, HET + vPMO-mScrAON versus HET + vPMO-mTtnAON and for PWd differences). No significant differences were observed among groups at day 0 and day 7. Data represent mean values ± SEM. Masson's trichrome staining of heart sections from HET animals injected with vPMO-mScrAONs and vPMO-mTtnAON. Scale bars, 1 mm.