Journal: Cell reports

Article Title: Opposing roles of p38α-mediated phosphorylation and PRMT1-mediated arginine methylation in driving TDP-43 proteinopathy.

doi: 10.1016/j.celrep.2024.115205

Figure Lengend Snippet: Figure 4. TDP-43 is methylated by PRMT1 at R293 (A) Sequence alignment of amino acids 287–307 of TDP-43 from diverse vertebrates. Conserved 292– 293 sites are bolded. S292 phosphorylation site and R293-G295 RGG-motif are highlighted in yellow and green, respectively. (B) Western blot of immunoprecipitated endoge- nous TDP-43 from SH-SY5Y-cells probed with antibodies against TDP-43, ADMA, MMA, and SDMA. TDP-43 with arginine methylation is marked by arrowheads. (C) Western blot of immunoprecipitated endoge- nous TDP-43 from SH-SY5Y-cells with siRNA- induced PRMT1 knockdown probed with anti- bodies against TDP-43 or MMA. A PRMT1 blot was run separately to confirm siRNA-mediated knockdown of PRMT1. (D) Western blot of expressed and immunopre- cipitated TDP-43WT using anti-FLAG antibody from SH-SY5Y-cells treated with DMSO (D) or with methyltransferase inhibitor AdOx (A) at a final concentration of 20 mM for 24 h. FL, full-length TDP-43; CTF35, C-terminal 35-kDa fragment of TDP-43. (E) Western blot of expressed and immunopre- cipitated TDP-43WT and TDP-43R293K using anti- FLAG antibody from SH-SY5Y cells probed with antibodies against TDP-43 and MMA. See also Figures S6 and S7.

Article Snippet: The generation of plasmids harboring N-terminally myc-tagged wild-type or M337V-mutant TDP-43 sequences were as described.53 Plasmids harboring C-terminally FLAG-tagged wild-type or mutant human TDP-43 and p38a sequences in pcDNA3.1+/C-(K)-DYK mammalian expression vector were purchased from Genscript and PRMT1 plasmid was purchased from Origene.

Techniques: Methylation, Sequencing, Phospho-proteomics, Western Blot, Immunoprecipitation, Knockdown, Concentration Assay

Journal: Cell reports

Article Title: Opposing roles of p38α-mediated phosphorylation and PRMT1-mediated arginine methylation in driving TDP-43 proteinopathy.

doi: 10.1016/j.celrep.2024.115205

Figure Lengend Snippet: Figure 6. Arginine methylation of TDP-43 regulates its aggregation (A) Western blot of total and pTDP-43 in RIPA and urea fractions of TDP-43WT-transfected SH-SY5Y-cells treated with DMSO (D) or with methyltransferase inhibitor AdOx (A) at a final concentration of 20 mM for 24 h. The positions of FLAG-tagged TDP-43 (exo) and endogenous TDP-43 (endo) are indicated. (B) Quantification of urea/RIPA ratio of total TDP-43 normalized to levels in DMSO-treated cells (mean ± SD, unpaired t test, n = 4). *p < 0.05. (C) Quantification of pTDP-43 in the urea fraction normalized to levels in DMSO-treated cells (mean ± SD, unpaired t test, n = 4). (D) Western blot of total and pTDP-43 in RIPA and urea fractions of TDP-43WT-transfected SH-SY5Y-cells co-transfected with empty control plasmid (Ctrl) or PRMT1. The positions of FLAG-tagged TDP-43 (exo) and endogenous TDP-43 (endo) are indicated. The positions of Myc-tagged PRMT1 (exo) and endogenous PRMT1 (endo) are also indicated. (E) Quantification of urea/RIPA ratio of total TDP-43 normalized to levels in Ctrl-plasmid co-transfected cells (mean ± SD, unpaired t test, n = 5). ***p < 0.001. (F) Quantification of pTDP-43 in the urea fraction normalized to levels in control cells (mean ± SD, unpaired t test, n = 5). ****p < 0.0001. See also Figure S7.

Article Snippet: The generation of plasmids harboring N-terminally myc-tagged wild-type or M337V-mutant TDP-43 sequences were as described.53 Plasmids harboring C-terminally FLAG-tagged wild-type or mutant human TDP-43 and p38a sequences in pcDNA3.1+/C-(K)-DYK mammalian expression vector were purchased from Genscript and PRMT1 plasmid was purchased from Origene.

Techniques: Methylation, Western Blot, Transfection, Concentration Assay, Control, Plasmid Preparation

Journal: Cell reports

Article Title: Opposing roles of p38α-mediated phosphorylation and PRMT1-mediated arginine methylation in driving TDP-43 proteinopathy.

doi: 10.1016/j.celrep.2024.115205

Figure Lengend Snippet: Figure 7. Crosstalk between PRMT1-catalyzed arginine methylation and p38a-mediated phosphorylation of TDP-43 (A) Western blot of immunoprecipitated FLAG-tagged TDP-43 from SH-SY5Y cells probed with antibodies against TDP-43, pTDP-43, and MMA. GAPDH serves as a loading control. The positions of FLAG-tagged TDP-43 (exo) and endogenous TDP-43 (endo) are indicated. (B) Western blot of immunoprecipitated FLAG-tagged TDP-43WT from SH-SY5Y cells with or without siRNA-induced p38a knockdown probed with antibodies against TDP-43, p38a, pTDP-43, and MMA. GAPDH serves as a loading control. (C) Quantification of TDP-43-CTF/full length-ratio (mean ± SD, unpaired t test, n = 3). **p < 0.01. (D) Quantification of mono-methylated TDP-43-CTF normalized to total TDP-43-CTF (mean ± SD, unpaired t test, n = 3). *p < 0.05. (E) Western blot of immunoprecipitated FLAG-tagged TDP-43WT from SH-SY5Y cells with or without PRMT1 overexpression probed with antibodies against TDP- 43, pTDP-43, and MMA. GAPDH serves as a loading control. The positions of FLAG-tagged TDP-43 (exo) and endogenous TDP-43 (endo) are indicated. The positions of Myc-tagged PRMT1 (exo) and endogenous PRMT1 (endo) are also indicated. See also Figure S7.

Article Snippet: The generation of plasmids harboring N-terminally myc-tagged wild-type or M337V-mutant TDP-43 sequences were as described.53 Plasmids harboring C-terminally FLAG-tagged wild-type or mutant human TDP-43 and p38a sequences in pcDNA3.1+/C-(K)-DYK mammalian expression vector were purchased from Genscript and PRMT1 plasmid was purchased from Origene.

Techniques: Methylation, Phospho-proteomics, Western Blot, Immunoprecipitation, Control, Knockdown, Over Expression

Journal: Nature Communications

Article Title: A super-enhancer-regulated RNA-binding protein cascade drives pancreatic cancer

doi: 10.1038/s41467-023-40798-6

Figure Lengend Snippet: a Venn diagram showing the overlap among eCLIP targets in AA0779E cells, genes downregulated upon HNRNPF knockdown in AA0779E cells, and genes downregulated upon Hnrnpf KO in mouse FC1245 cells. b Browser tracks displaying hnRNP F binding to PRMT1 mRNA; 3’UTR is boxed in red. c RT-qPCR showing expression levels of HNRNPF and PRMT1 , normalized to GAPDH , in MIA PaCa-2 cells knocked down with siRNA against HNRNPF or a non-targeting control ( n = 3 biological replicates). d Representative immunoblot (left) and quantification (right) showing Prmt1 levels in FC1245 parental and Hnrnpf KO cells ( n = 3 biological replicates). e RT-qPCR showing levels of an exogenously expressed luciferase gene containing the 3’UTR of Prmt1 , normalized to RFP driven by an independent promoter on the same plasmid, in FC1245 parental, Hnrnpf KO, or Hnrnpf KO cells rescued by exogenous HNRNPF re-expression ( n = 6 biological replicates). f Representative immunoblot from two independent experiments showing Prmt1 and hnRNP F levels in FC1245 parental, Hnrnpf KO, or Hnrnpf KO cells rescued with re-expression of exogenous Prmt1. g Cell confluence determined using IncuCyte software from phase-contrast images of FC1245 parental, Hnrnpf KO, or Hnrnpf KO cells rescued with re-expression of exogenous Prmt1 ( n = 3 independent experiments). Data represent means ± SEM in ( c – e ) and ( g ). Unpaired two-tailed t-test with two-stage step-up multiple comparison correction was performed in ( c ), one-way ANOVA followed by Tukey’s multiple comparisons test in ( d , e ), and one-way ANOVA with Sidak’s multiple comparisons test in ( g ). ns: not significant. Source data are provided as a Source Data file.

Article Snippet: The following antibodies were utilized: human hnRNP F (Santa Cruz Biotechnology Cat# sc-32309, RRID:AB_627732, 1:1000), mouse/human hnRNP F (Abcam Cat# ab50982, RRID:AB_880477, 1:1000), PRMT1 (R and D Systems Cat# AF6016, RRID:AB_1964684, 1:1000), Vinculin (R and D Systems Cat# MAB6896, RRID:AB_10992930, 1:1000), human UBAP2L (Bethyl Cat# A300-533A, RRID:AB_477953, 1:10,000), mouse/human UBAP2L (Bethyl Cat# A300-534A, RRID:AB_2272582, 1:5000), Anti-dimethyl-Arginine Antibody, asymmetric (Millipore, at# 07-414, RRID:AB_310596, 1:1000), β -Actin D6A8 (Cell Signaling Technology Cat# 8457, RRID:AB_10950489 1:1000), RPL31 (ABclonal Cat# A17527, RRID:AB_2772081, 1:1000), EEF1D (ABclonal Cat# A2509, RRID:AB_2764400, 1:1000), anti-rabbit IgG-HRP (Santa Cruz Biotechnology Cat# sc-2004, RRID:AB_631746, 1:10,000), anti-mouse IgG-HRP (Santa Cruz Biotechnology Cat# sc-2005, RRID:AB_631736, 1:10,000), anti-goat IgG-HRP (Millipore Cat# 401515-2ML, RRID:AB_10682600, 1:10,000).

Techniques: Knockdown, Binding Assay, Quantitative RT-PCR, Expressing, Control, Western Blot, Luciferase, Plasmid Preparation, Software, Two Tailed Test, Comparison

Journal: Nature Communications

Article Title: A super-enhancer-regulated RNA-binding protein cascade drives pancreatic cancer

doi: 10.1038/s41467-023-40798-6

Figure Lengend Snippet: a Representative immunoblot from two independent experiments of Prmt1 levels in FC1245 parental, Prmt1 KO, or Prmt1 KO cells rescued with re-expression of exogenous Prmt1. Cell confluence determined using IncuCyte software from phase-contrast images of b FC1245 parental, Prmt1 KO, or Prmt1 KO cells rescued with re-expression of exogenous wild-type Prmt1 ( n = 3 independent experiments) and c FC1245 parental, Prmt1 KO, or Prmt1 KO cells expressing exogenous catalytically dead Prmt1 ( n = 4 independent experiments). d Tumor weights from C57BL/6J mice orthotopically transplanted with FC1245 parental ( n = 3), Prmt1 KO ( n = 4), or Prmt1 KO cells re-expressing Prmt1 ( n = 4) and sacrificed 4 weeks after transplant. Data represent means ± SD in ( b , c ) and means ± SEM in ( d ). One-way ANOVA with Sidak’s multiple comparisons test was performed in ( b , c ) and one-way ANOVA followed by Tukey’s multiple comparison test in ( d ). ns: not significant. Source data are provided as a Source Data file.

Article Snippet: The following antibodies were utilized: human hnRNP F (Santa Cruz Biotechnology Cat# sc-32309, RRID:AB_627732, 1:1000), mouse/human hnRNP F (Abcam Cat# ab50982, RRID:AB_880477, 1:1000), PRMT1 (R and D Systems Cat# AF6016, RRID:AB_1964684, 1:1000), Vinculin (R and D Systems Cat# MAB6896, RRID:AB_10992930, 1:1000), human UBAP2L (Bethyl Cat# A300-533A, RRID:AB_477953, 1:10,000), mouse/human UBAP2L (Bethyl Cat# A300-534A, RRID:AB_2272582, 1:5000), Anti-dimethyl-Arginine Antibody, asymmetric (Millipore, at# 07-414, RRID:AB_310596, 1:1000), β -Actin D6A8 (Cell Signaling Technology Cat# 8457, RRID:AB_10950489 1:1000), RPL31 (ABclonal Cat# A17527, RRID:AB_2772081, 1:1000), EEF1D (ABclonal Cat# A2509, RRID:AB_2764400, 1:1000), anti-rabbit IgG-HRP (Santa Cruz Biotechnology Cat# sc-2004, RRID:AB_631746, 1:10,000), anti-mouse IgG-HRP (Santa Cruz Biotechnology Cat# sc-2005, RRID:AB_631736, 1:10,000), anti-goat IgG-HRP (Millipore Cat# 401515-2ML, RRID:AB_10682600, 1:10,000).

Techniques: Western Blot, Expressing, Software, Comparison

Journal: Nature Communications

Article Title: A super-enhancer-regulated RNA-binding protein cascade drives pancreatic cancer

doi: 10.1038/s41467-023-40798-6

Figure Lengend Snippet: a Representative immunoblot (top) and densitometric quantification (bottom) of whole cell extracts from puromycin-treated FC1245 parental, Prmt1 KO, or Prmt1 KO cells rescued by re-expression of exogenous Prmt1 ( n = 5 independent experiments). b Polysome profiling of FC1245 parental or Prmt1 KO cells showing the normalized absorbance at 260 nm during fractionation of polysomes ( n = 3 biological replicates). c Representative immunoblot from three independent experiments showing Prmt1 and asymmetrically dimethylated arginine-containing protein (ASYM24) levels in FC1245 parental and Prmt1 KO cells. d (Top) Schematic of UBAP2L protein structure displaying the N-terminal ubiquitin-associated (UBA) domain and the Arg–Gly–Gly (RGG) domain that contains the two Arginines asymmetrically dimethylated by Prmt1 (red ovals). (Bottom) Extracted MS1 chromatographic peak of the listed Ubap2l peptide, showing dimethylation at both R’s in the N-terminal RGGR motif. FC1245 parental peptide was labeled light and the Prmt1 KO sample was labeled heavy. e Representative images (left) and quantification (right) of UBAP2L IHC from a human PDAC tissue microarray containing biological replicates of early ( n = 149) or late ( n = 13) stage PDAC. Scale bar: 100 µm. f Representative immunoblot (left) and quantification (right) of input and RNA interactome capture (RIC) fractions using antibodies against Ubap2l ( n = 7 independent experiments) and Rps3 ( n = 3 independent experiments). Data represent the mean ± SEM in ( a , b ) and ( f ). Box plots indicate median (middle line), 25th, 75th percentile (box), 10th and 90th percentile (whiskers), and outliers (single points). One-way ANOVA followed by Dunn’s multiple comparisons test was performed in ( a ), unpaired two-tailed t-test in ( b ) and ( f ), and two-tailed Mann–Whitney test in ( e ). ns: not significant. Source data are provided as a Source Data file.

Article Snippet: The following antibodies were utilized: human hnRNP F (Santa Cruz Biotechnology Cat# sc-32309, RRID:AB_627732, 1:1000), mouse/human hnRNP F (Abcam Cat# ab50982, RRID:AB_880477, 1:1000), PRMT1 (R and D Systems Cat# AF6016, RRID:AB_1964684, 1:1000), Vinculin (R and D Systems Cat# MAB6896, RRID:AB_10992930, 1:1000), human UBAP2L (Bethyl Cat# A300-533A, RRID:AB_477953, 1:10,000), mouse/human UBAP2L (Bethyl Cat# A300-534A, RRID:AB_2272582, 1:5000), Anti-dimethyl-Arginine Antibody, asymmetric (Millipore, at# 07-414, RRID:AB_310596, 1:1000), β -Actin D6A8 (Cell Signaling Technology Cat# 8457, RRID:AB_10950489 1:1000), RPL31 (ABclonal Cat# A17527, RRID:AB_2772081, 1:1000), EEF1D (ABclonal Cat# A2509, RRID:AB_2764400, 1:1000), anti-rabbit IgG-HRP (Santa Cruz Biotechnology Cat# sc-2004, RRID:AB_631746, 1:10,000), anti-mouse IgG-HRP (Santa Cruz Biotechnology Cat# sc-2005, RRID:AB_631736, 1:10,000), anti-goat IgG-HRP (Millipore Cat# 401515-2ML, RRID:AB_10682600, 1:10,000).

Techniques: Western Blot, Expressing, Fractionation, Ubiquitin Proteomics, Labeling, Microarray, Two Tailed Test, MANN-WHITNEY

Journal: Nature Communications

Article Title: A super-enhancer-regulated RNA-binding protein cascade drives pancreatic cancer

doi: 10.1038/s41467-023-40798-6

Figure Lengend Snippet: a Representative anti-puromycin immunoblot (top) and densitometric analysis (bottom) of extracts from puromycin-treated MIA PaCa-2 cells knocked down using siRNAs against UBAP2L , PRMT1 , or a non-targeting control ( n = 3 biological replicates). b Representative immunoblot from two independent experiments showing Ubap2l levels in FC1245 parental and Ubap2l KO cells. c Cell confluence determined using IncuCyte software from phase-contrast images of FC1245 parental or Ubap2l KO cells ( n = 3 independent experiments). d mRNA expression of the indicated rRNAs, normalized to Gapdh , in FC1245 parental, Prmt1 KO, and Ubap2l KO cells ( n = 3 biological replicates). e Representative immunoblot (top) and quantification (bottom) showing Rpl31 protein levels, normalized to Vinculin, in FC1245 parental and Ubap2l KO cells ( n = 3). f Tumor weights from mice orthotopically transplanted with FC1245 parental or Ubap2l KO cells ( n = 3). g Representative images (left) and quantification (right) of tumor sections of FC1245 parental and Ubap2l KO tumors stained with Rpl31 ( n = 6 from 2 fields per sample; scale bar: 100 μm). Data represent means ± SEM in ( a , d , e – g ), and means ± SD in ( c ). One-way ANOVA followed by Tukey’s multiple comparison test was performed in ( a ), unpaired two-tailed t-test in ( c ) and ( e – g ), and two-way ANOVA with Dunnett’s multiple comparison test in ( d ). ns: not significant. Source data are provided as a Source Data file.

Article Snippet: The following antibodies were utilized: human hnRNP F (Santa Cruz Biotechnology Cat# sc-32309, RRID:AB_627732, 1:1000), mouse/human hnRNP F (Abcam Cat# ab50982, RRID:AB_880477, 1:1000), PRMT1 (R and D Systems Cat# AF6016, RRID:AB_1964684, 1:1000), Vinculin (R and D Systems Cat# MAB6896, RRID:AB_10992930, 1:1000), human UBAP2L (Bethyl Cat# A300-533A, RRID:AB_477953, 1:10,000), mouse/human UBAP2L (Bethyl Cat# A300-534A, RRID:AB_2272582, 1:5000), Anti-dimethyl-Arginine Antibody, asymmetric (Millipore, at# 07-414, RRID:AB_310596, 1:1000), β -Actin D6A8 (Cell Signaling Technology Cat# 8457, RRID:AB_10950489 1:1000), RPL31 (ABclonal Cat# A17527, RRID:AB_2772081, 1:1000), EEF1D (ABclonal Cat# A2509, RRID:AB_2764400, 1:1000), anti-rabbit IgG-HRP (Santa Cruz Biotechnology Cat# sc-2004, RRID:AB_631746, 1:10,000), anti-mouse IgG-HRP (Santa Cruz Biotechnology Cat# sc-2005, RRID:AB_631736, 1:10,000), anti-goat IgG-HRP (Millipore Cat# 401515-2ML, RRID:AB_10682600, 1:10,000).

Techniques: Western Blot, Control, Software, Expressing, Staining, Comparison, Two Tailed Test

Journal: Nature Communications

Article Title: A super-enhancer-regulated RNA-binding protein cascade drives pancreatic cancer

doi: 10.1038/s41467-023-40798-6

Figure Lengend Snippet: a Genome browser tracks (from one of two independent experiments) showing Myc binding at the Hnrnpf , Prmt1 , and Ubap2l loci upon activation of MycER T2 with 4-OHT for the indicated time points. b RT-qPCR showing Hnrnpf, Prmt1 , and Ubap2l expression, normalized to Actb , in mouse pancreatic epithelial cells in which MycER T2 was activated with 4-OHT for the indicated time points. Data represent the mean ± SEM from three biological replicates. One-way ANOVA followed by Tukey’s multiple comparison test was used. ns: not significant. Source data are provided as a Source Data file.

Article Snippet: The following antibodies were utilized: human hnRNP F (Santa Cruz Biotechnology Cat# sc-32309, RRID:AB_627732, 1:1000), mouse/human hnRNP F (Abcam Cat# ab50982, RRID:AB_880477, 1:1000), PRMT1 (R and D Systems Cat# AF6016, RRID:AB_1964684, 1:1000), Vinculin (R and D Systems Cat# MAB6896, RRID:AB_10992930, 1:1000), human UBAP2L (Bethyl Cat# A300-533A, RRID:AB_477953, 1:10,000), mouse/human UBAP2L (Bethyl Cat# A300-534A, RRID:AB_2272582, 1:5000), Anti-dimethyl-Arginine Antibody, asymmetric (Millipore, at# 07-414, RRID:AB_310596, 1:1000), β -Actin D6A8 (Cell Signaling Technology Cat# 8457, RRID:AB_10950489 1:1000), RPL31 (ABclonal Cat# A17527, RRID:AB_2772081, 1:1000), EEF1D (ABclonal Cat# A2509, RRID:AB_2764400, 1:1000), anti-rabbit IgG-HRP (Santa Cruz Biotechnology Cat# sc-2004, RRID:AB_631746, 1:10,000), anti-mouse IgG-HRP (Santa Cruz Biotechnology Cat# sc-2005, RRID:AB_631736, 1:10,000), anti-goat IgG-HRP (Millipore Cat# 401515-2ML, RRID:AB_10682600, 1:10,000).

Techniques: Binding Assay, Activation Assay, Quantitative RT-PCR, Expressing, Comparison

Journal: Nature Communications

Article Title: A super-enhancer-regulated RNA-binding protein cascade drives pancreatic cancer

doi: 10.1038/s41467-023-40798-6

Figure Lengend Snippet: a Caspase-3/7 activity assay showing quantification of % apoptotic cells (left) and representative images (right) of MIA-PaCa2 cells treated with the indicated TC-E 5003 doses for 16 h ( n = 2 independent experiments). The cells labeled in green are undergoing apoptosis. Scale bar: 100 μm. b Cell viability assay showing dose-response curves for MYC-high organoids hF3 ( N = 4), hF23 ( N = 3), and hT3 ( N = 3) and MYC-low organoids hM19A ( N = 4), hM1E ( N = 4), and hF44 ( N = 4) treated with the indicated PRMT1 inhibitors for 5 days. c Tumor weights, normalized to vehicle, from mice orthotopically transplanted with FC1245 parental cells and treated 10 days post-transplant with AMI-408 or vehicle ( n = 4) or with TC-E 5003 or vehicle ( n = 7) for 2 weeks. d Representative images (top) and quantification (bottom) of Ki67-stained tumor sections from mice treated with vehicle or AMI-408 ( N = 16 from 4 fields per sample) or with vehicle or TC-E 5003 ( N = 14 from 2 fields per sample). Scale bar: 100 µm. e Model showing that the HNRNPF SE regulates tumor growth by driving the expression of HNRNPF . In turn, hnRNP F regulates Prmt1 mRNA levels. Prmt1 asymmetrically dimethylates Ubap2l, which regulates the expression of rRNA and ribosomal proteins to control protein translation. Increased protein translation leads to increased tumor growth. Myc transcriptionally coordinates this entire program. The figure was partly generated using Servier Medical Art, provided by Servier, licensed under a Creative Commons Attribution 3.0 unported license. Data represent the mean ± SEM in ( a – c ). Data in ( a ) represent means and the range. Box plots in ( d ) indicate median (middle line), 25th, 75th percentile (box), minima and maxima (whiskers). One-way ANOVA with Tukey’s multiple comparison test was used to compare IC50s in ( b ). For both PRMT1-2e and TC-E 5003, hM19A vs hF3, hF23 or hT3 p < 0.0001, hM1E vs hF3, hF23 or hT3 p < 0.0001, hF44 vs hF3, hF23 or hT3 p < 0.0001. The remaining comparisons were not significant. An unpaired two-tailed t-test was performed in ( c ) and a two-tailed Mann–Whitney test in ( d ). ns: not significant. Source data are provided as a Source Data file.

Article Snippet: The following antibodies were utilized: human hnRNP F (Santa Cruz Biotechnology Cat# sc-32309, RRID:AB_627732, 1:1000), mouse/human hnRNP F (Abcam Cat# ab50982, RRID:AB_880477, 1:1000), PRMT1 (R and D Systems Cat# AF6016, RRID:AB_1964684, 1:1000), Vinculin (R and D Systems Cat# MAB6896, RRID:AB_10992930, 1:1000), human UBAP2L (Bethyl Cat# A300-533A, RRID:AB_477953, 1:10,000), mouse/human UBAP2L (Bethyl Cat# A300-534A, RRID:AB_2272582, 1:5000), Anti-dimethyl-Arginine Antibody, asymmetric (Millipore, at# 07-414, RRID:AB_310596, 1:1000), β -Actin D6A8 (Cell Signaling Technology Cat# 8457, RRID:AB_10950489 1:1000), RPL31 (ABclonal Cat# A17527, RRID:AB_2772081, 1:1000), EEF1D (ABclonal Cat# A2509, RRID:AB_2764400, 1:1000), anti-rabbit IgG-HRP (Santa Cruz Biotechnology Cat# sc-2004, RRID:AB_631746, 1:10,000), anti-mouse IgG-HRP (Santa Cruz Biotechnology Cat# sc-2005, RRID:AB_631736, 1:10,000), anti-goat IgG-HRP (Millipore Cat# 401515-2ML, RRID:AB_10682600, 1:10,000).

Techniques: Activity Assay, Labeling, Viability Assay, Staining, Expressing, Control, Generated, Comparison, Two Tailed Test, MANN-WHITNEY