Fig. 4: HMGB1-modulated TRIM30α to induce senescence.

HMGB1-WT and -KO MEFs were treated with 100 ng/ml Dox for 3 days, then western blotting (A) and qPCR (B) were performed to assess TRIM30α expression. Cells were transfected with Si C or Si HMGB1 24 h prior to Dox treatment to B16-F10 cells for 3 days, and immunoprecipitation was performed with anti-HMGB1 antibodies. PCR was carried out using TRIM30α promoter-specific primers for ChIP assays (C), and the binding level was quantified (D). TRIM30α promoter activity was measured by luciferase assay at day 3 after Dox treatment (E). Cells were transfected with Si HMGB1 in a concentration-dependent manner for 2 days, and western blotting was performed (F). Cells were transfected with HMGB1 plasmid (G) or Si HMGB1 (H) for 2 days, and western blotting was performed. HMGB1-KO MEFs were transfected with 100 nM Si C or Si TRIM30α for 3 days. Morphological changes were photographed (I), protein expression was evaluated by western blotting (J), and mRNA expression was estimated by qPCR (K). Quantitative data are represented as mean ± S.D., *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, n = 3 independent trials Scale bars, 50 μm.