Journal: Genome Biology
Article Title: O-GlcNAcylation of NONO mediates alternative splicing of SETMAR and facilitates NHEJ repair
doi: 10.1186/s13059-026-03930-5
Figure Lengend Snippet: O-GlcNAcylation of NONO promotes radioresistance in hepatocellular carcinoma. A Analysis of NONO O-GlcNAcylation was conducted in human liver epithelial cells (THLE2) and hepatocellular carcinoma cells (HepG2, HCCLM9, Huh7) usingIP/Western blotting with indicated antibodies. B Generation of stable HCCLM9 cells with NONO knockdown. Western blot analysis was performed to assess the efficiency of NONO knockdown using two shRNA constructs(shNONO #1 and shNONO #2). GAPDH was used as a loading control. C Upper panel: O-GlcNAcylation at Ser147 promotes cell survival post-IR treatment. NONO-knockdown HCCLM9 cells were transfected with SFB-NONO WT or S147A mutant and subjected to clonogenic survival assays post-IR treatment. Cells were treated with the indicated doses of IR and further incubated for 7–10 days. Lower panel: Quantitative analysis of clonogenic survival assays. D Schematic diagram illustrating the radiotherapy process for NTG mice. Mice injected with control or NONO knockdown cells, as well as cells reconstituted with NONO WT or S147A and wereexposed to 8 Gy of IR twice. E O-GlcNAcylation of NONO enhances radioresistance. Mice were subcutaneously injected with 7 × 10 6 control or NONO knockdown cells, or cells reconstituted with NONO WT or S147A, and exposed to 8 Gy of IR twice or not, when tumors reached a similar size (about 100 mm. 3 ). Representative images of xenograft tumors are shown ( n = 6/group). F Quantification analysis of xenograft tumor volumes from ( E ). Data are presented as mean ± SD. Statistical significance was determined using one-way ANOVA. ** P < 0.01, *** P < 0.001. G Hematoxylin and eosin (H&E) staining and IHC analysis of NONO and H3K36me2 in xenograft tumors were performed, comparing controland NONO knockdown groups, as well as tumors reconstituted with NONO WT or S147A ( n = 6). Scale bar, 30 μm. H Quantification of H3K36me2 levels from ( G ). Data are presented as mean ± SD. Statistical significance was determined using one-way ANOVA. ** P < 0.01, *** P < 0.001
Article Snippet: The HEK293T, U2OS, THLE2, Huh7, HepG2 and HCCLM9 cell lines were purchased from American Type Cell Culture (ATCC), and cultured in Dulbecco’s modified Eagle medium (Gibco) supplemented with 10% fetal bovine serum (FBS) (Gibco), 100 U/mL penicillin, and 100 μg/mL streptomycin in the presence of 5% CO 2 (v/v) at 37 °C.
Techniques: Western Blot, Knockdown, shRNA, Construct, Control, Transfection, Mutagenesis, Incubation, Injection, Staining
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