Fig. 3: Cisplatin (CP) induced SET8 and H4K20me1 expression, caspase 3 cleavage, and co-localization of SET8 and PTEN in the nucleus of TKPTs.

Mouse proximal tubule cells (TKPTs) were exposed to various concentrations (0, 5, 10, and 20 μM) of CP for 24 h (A, B) or treated with cisplatin (20 μM) for 0, 6, 12, and 24 h (C, D). Cell lysates were prepared and subjected to immunoblot analysis against SET8, H4K20me1, histone H4 (H4), cleaved cas3, PTEN, Tubulin, or GAPDH. The expression levels of all those proteins were quantified by densitometry. SET8, cleaved cas3, and PTEN in Fig. 3A were normalized by Tubulin (B), and those protein levels in Fig. 3C were normalized by GAPDH (D); H4K20me1 in Fig. 3A, C, was normalized by H4 (B, D). IF staining for SET8 and PETN is indicated (E). DAPI staining was used to localize nuclei in TKPT. Scale bars = 20 μm. Arrows: nuclei with SET8 and PETN staining. F Western blotting and quantification (G) of the nuclear and cytoplasmic distribution of SET8 and PTEN at various concentrations of cisplatin. Histone H3 (H3) and Tubulin serve as nuclear and cytoplasmic markers, respectively. Data are represented as the mean ± SEM of at least three experiments. *P < 0.05, **P < 0.01, ***P < 0.001.