Fig. 1: Deficiency of TET2 promotes YAP activity and sensitizes HCC cells to sorafenib and verteporfin.

Deficiency of TET2 sensitizes HCC cells to sorafenib. Control and TET2 KO HepG2 cells were treated with different concentration of sorafenib as indicated for three days (A), or 10 μM sorafenib for different times as indicated (B). Cell viability was analyzed in top panels. Western blot analysis of cleaved PARP was performed in bottom panels. C TET2 deficiency sensitizes tumor cells to sorafenib treatment in vivo. Control and TET2 KO MHCC97H cells were subcutaneously injected into the left flanks of athymic nude mice (n = 6 per group). Tumor inhibition rate of sorafenib was measured and calculated every other day as indicated. D Western blot analysis of tumors from nude mice was performed with indicated antibodies. Phosphorylation level of YAP Ser127 is significantly downregulated in sgTET2 HCC cells (E) and livers of Tet2 knockout mice (F). G Western blot analysis of cytoplasm and nuclear extracts derived from Control and TET2 KO cell lines (HepG2 or MHCC97H). H Immunofluorescence of YAP localization in Control or TET2 KO HepG2 cells. YAP target genes CTGF and CYR61 are distinctly upregulated in sgTET2 HCC cells (I) and livers of Tet2 knockout mice (J). K Re-introduction of WT TET2 not catalytic mutant TET2 (R1896S) can rescue phosphorylation level of YAP Ser127 in sgTET2 HepG2. L Reintroduction of WT TET2 can rescue mRNA expression of CTGF and CYR61. M A total of 253 LIHC tumors from TCGA database were divided into two groups based on TET2 mRNA levels (top and bottom 50% TET2 expression), and their relative YAP activities were qualified and plotted as described in the Methods. LIHC, liver hepatocellular carcinoma. N, O Deficiency of TET2 sensitizes HCC cells to verteporfin. Control and TET2 KO HepG2 cells were treated with different concentration of verteporfin as indicated for two days (N), or 1 μM verteporfin for different times as indicated (O). Cell viability was analyzed in top panels. Western blot analysis of cleaved PARP was performed in bottom panels. Data are presented as mean ± s.d., n = 3 independent repeats. Unpaired, two-tailed t-test.