Journal: Scientific reports

Article Title: Arginine methylation of hnRNPUL1 regulates interaction with NBS1 and recruitment to sites of DNA damage.

doi: 10.1038/srep10475

Figure Lengend Snippet: Figure 1. The RGG/RG motif of hnRNPUL1 is methylated by PRMT1 in vitro. A) SAP designates the SAF- A/B, Acinus and PIAS motif, while BBS denotes BRD7-binding site. The RGG/RG motifs located from 609 to 658 are shown. The RG and RGG repeats are bold and underlined. B) The human hnRNPUL1 peptide sequences fused to recombinant GST used for the methylation assay of panel C. C) PRMT1 in vitro methylation assay using GST-hnRNPUL1 proteins of panel B with H3-SAM. Proteins were resolved by SDS- PAGE, stained with Coomassie blue (left panel), dried and visualized by fluorography (right panel). GST- MRE11:RGG (glycine arginine rich) and GST were used as the positive and negative control, respectively. The indicated sequences (RGG or RG) fused to GST migrate at a similar molecular mass as GST alone because of the short added sequence. GST-MRE11:RGG harbors an extra ~60aa and migrates close to 33 kDa. The migration of GST-PRMT1 is shown with arrows.

Article Snippet: The human hnRNPUL1 cDNA purchased from ORIGENE (Rockville, MD) was FLAG epitope-tagged and subcloned into pcDNA3.1 with the following primers 5′ -GGG GGA TCC GAT GTG CGC CGT CTG AAG GTG-3′ and 5′ -GGG GTC GAC CTA CTG TGT ACT TGT GCC ACC3.

Techniques: Methylation, In Vitro, Binding Assay, Recombinant, SDS Page, Staining, Negative Control, Sequencing, Migration

Journal: Scientific reports

Article Title: Arginine methylation of hnRNPUL1 regulates interaction with NBS1 and recruitment to sites of DNA damage.

doi: 10.1038/srep10475

Figure Lengend Snippet: Figure 2. The RGG/RG motif of hnRNPUL1 is methylated by PRMT1 in vivo via a physical association. A) Whole cell lysates from HEK293 cells transfected with siControl (CTL) and siPRMT1 were subjected to immunoprecipitation (IP) 48 h post-transfection with the anti-hnRNPUL1 antibody. Whole cell lysates (WCL) and immunoprecipitants were immunoblotted with anti-asymmetrical dimethylarginines (ASYM25b), anti-hnRNPUL1, anti-PRMT1 and anti-Tubulin antibodies. B) HEK293 cells transfected with wild type FLAG-hnRNPUL1 and FLAG-hnRNPUL1RK were lysed and anti-FLAG immunoprecipitations were performed. The bound proteins were separated by SDS-PAGE and immunoblotted with the indicated antibodies. Tubulin was used as loading control.

Article Snippet: The human hnRNPUL1 cDNA purchased from ORIGENE (Rockville, MD) was FLAG epitope-tagged and subcloned into pcDNA3.1 with the following primers 5′ -GGG GGA TCC GAT GTG CGC CGT CTG AAG GTG-3′ and 5′ -GGG GTC GAC CTA CTG TGT ACT TGT GCC ACC3.

Techniques: Methylation, In Vivo, Transfection, Immunoprecipitation, SDS Page, Control

Journal: Scientific reports

Article Title: Arginine methylation of hnRNPUL1 regulates interaction with NBS1 and recruitment to sites of DNA damage.

doi: 10.1038/srep10475

Figure Lengend Snippet: Figure 3. Methylation of hnRNPUL1 is required for its interaction with NBS1. A) U2OS cells were transfected with empty vector (pcDNA 3.1), FLAG-hnRNPUL1 wild type, FLAG-hnRNPUL1RK mutant and YFP-NBS1. Whole cell lysates (WCL) and FLAG-immunoprecipitants were immunoblotted with the indicated antibodies. B) HEK293 cells were transfected with siControl and siPRMT1 along with YFP-NBS1 and FLAG-hnRNPUL1. The whole cell lysates (WCL) and FLAG immunoprecipitates were subjected to western blot analysis using the antibodies indicated.

Article Snippet: The human hnRNPUL1 cDNA purchased from ORIGENE (Rockville, MD) was FLAG epitope-tagged and subcloned into pcDNA3.1 with the following primers 5′ -GGG GGA TCC GAT GTG CGC CGT CTG AAG GTG-3′ and 5′ -GGG GTC GAC CTA CTG TGT ACT TGT GCC ACC3.

Techniques: Methylation, Transfection, Plasmid Preparation, Mutagenesis, Western Blot

Journal: Scientific reports

Article Title: Arginine methylation of hnRNPUL1 regulates interaction with NBS1 and recruitment to sites of DNA damage.

doi: 10.1038/srep10475

Figure Lengend Snippet: Figure 4. RGG/RG motif regulates GFP-hnRNPUL1 recruitment at breaks. A) U2OS cells were transfected with wild type GFP-hnRNPUL1 and GFP-hnRNPUL1RK and subjected to laser micro-irradiation treatment (forms a DNA damage ‘track’; white box). Exclusion of GFP-UL1 proteins from laser damage sites (2 min post-laser scissor damage) was monitored via live imaging (exclusion) or recruitment in the presence of DRB (inclusion) B) Quantification of exclusion and inclusion patterns of GFP-UL1 proteins are indicated as a percentage.

Article Snippet: The human hnRNPUL1 cDNA purchased from ORIGENE (Rockville, MD) was FLAG epitope-tagged and subcloned into pcDNA3.1 with the following primers 5′ -GGG GGA TCC GAT GTG CGC CGT CTG AAG GTG-3′ and 5′ -GGG GTC GAC CTA CTG TGT ACT TGT GCC ACC3.

Techniques: Transfection, Irradiation, Imaging

Journal: Discover oncology

Article Title: LINC00461 promotes bladder cancer cells EMT through miR-518b/HNRNPUL1 axis.

doi: 10.1007/s12672-024-01294-5

Figure Lengend Snippet: Fig. 3 LINC00461 positively regulates HNRNPUL1 expression via sponging miR-518b. A Starbase predicted binding sites between LINC00461 and miR-518b. B Dua-luciferase reporter assay validated interaction between LINC00461 and miR-518b. C The expression of miR- 518b in BC tissues and normal tissues by qRT-PCR (n = 32). D Pearson correlation analysis of the correlation between the relative expression levels of miR-518b and LINC00461. E Starbase predicted binding sites between HNRNPUL1-wild type or HNRNPUL1-mutant type and miR- 518b. F Dua-luciferase reporter assay validated interaction between miR-518b and HNRNPUL1. G The inhibitory efficiency of the miR-518b inhibitor was demonstrated by means of qRT-PCR. H, I Detection of the expression of HNRNPUL1 in BC cells transfected with miR-518b inhibitor through qRT-PCR and Western blotting. J, K Detection of the expression of HNRNPUL1 in BC cells transfected with miR-518b mimic through qRT-PCR and Western blotting. L Protein levels of HNRNPUL1 in cells transfected with siLINC00461 were confirmed by Western blot- ting. *p < 0.05

Article Snippet: The primary antibodies included HNRNPUL1 (Cat. No: 10578-1-AP) and β-actin (Cat. No: 81115-1-RR) were purchased from Proteintech Group (Wuhan, China).

Techniques: Expressing, Binding Assay, Luciferase, Reporter Assay, Quantitative RT-PCR, Mutagenesis, Transfection, Western Blot

Journal: Discover oncology

Article Title: LINC00461 promotes bladder cancer cells EMT through miR-518b/HNRNPUL1 axis.

doi: 10.1007/s12672-024-01294-5

Figure Lengend Snippet: Fig. 4 Knockdown of LINC00461 inhibits BC cells EMT through miR-518b/HNRNPUL1 axis. A The miR-518b inhibitor partially reversed cell proliferation inhibition induced by silencing LINC00461. B The miR-518b inhibitor partially reversed cell invasion induced by silencing LINC00461 (scale bar, 10 μm). C The miR-518b inhibitor partially reversed the inhibition of EMT and HNRNPUL1 protein expression induced by silencing LINC00461. *p < 0.05, **p < 0.01

Article Snippet: The primary antibodies included HNRNPUL1 (Cat. No: 10578-1-AP) and β-actin (Cat. No: 81115-1-RR) were purchased from Proteintech Group (Wuhan, China).

Techniques: Knockdown, Inhibition, Expressing