FIGURE 6.

Differential regulation of p63 expression by Gli1 and Gli2. A and B, mouse PEB cells were transduced with control vector, Gli3T, shRNA-Gli1 (shGli1), or shRNA-Gli2 (shGli2) by lentiviral infection. Cell lysates were collected and subjected to Western blotting analysis for expression of p63, Gli1, Gli2, Gli3T, and Erk2 (A) or quantitative RT-PCR for the expression of ΔNp63 and GAPDH (B). The vector group was normalized to GAPDH and set at 1. C, endogenous Gli1, 2, and 3 binding to the p63 promoter in PEB cells was examined by chromatin immunoprecipitation as described under “Experimental Procedures.” D–F, he primary prostate cells were transduced with control vector (Fu-CRW), Gli3T, shRNA-Gli1, or shRNA-Gli2 by lentiviral infection. After 5 days of growing in Matrigel, RFP was expressed in the transduced cells. RFP⁺ cells were sorted and remixed with Matrigel and subjected to the sphere formation assay. The number of primary (D) and secondary spheres (E) was recorded, and images of the spheres were taken (F). Scale bars = 200 μm. *, p < 0.05; **, p < 0.01; ***, p < 0.001; N.S., no significance (two-tailed Student t test).