Figure 8.

UNC119Ab and KRASB WT synergistically enhance p53 expression, which depends on RASSF6. KRAS silencing and RASSF6 silencing were performed in HCT116 cells as described for Fig. 7. GFP-UNC119Ab and Myc2-KRASB WT were expressed, and endogenous p53 was immunostained. Nuclear p53 signal was measured by ImageJ in the indicated number of cells. A, UNC119Ab augmented the signal of p53 in the nucleus (siCont), but the silencing of KRAS and RASSF6 reduced it. B, KRASB WT further enhanced the nuclear p53 signal in UNC119Ab-expressing cells (UNC119Ab+KRASB WT, siCont), whereas RASSF6 silencing abolished the effect. C, HCT116 cells were infected with retrovirus vectors harboring GFP or GFP-UNC119Ab and cultured in the medium containing 1 mg/liter of puromycin. 2 days later, the silencing of KRAS and RASSF6 was performed. 72 h later, whole-cell lysates were immunoblotted with the indicated antibodies. White and black arrows indicate GFP-UNC119Ab and its degraded product. The efficiency of knockdown was evaluated for KRAS by Western blotting and for RASSF6 by quantitative RT-PCR. The data are means with S.E. *, p < 0.05; ***, p < 0.001; n.s, not significant. Bar, 10 μm. Three experiments were performed. Numbers of samples are shown. Statistical analyses were performed with one-way ANOVA with Tukey's test.