FIG. 4.

Repression of mTOR/S6 kinase signaling is responsible for PTEN-mediated repression of RNA Pol I-dependent transcription. (A) Decreasing PTEN expression does not affect RNA Pol I-dependent transcription in cells treated with the mTOR inhibitor rapamycin. LN18 cells were transiently transfected with PrHuCAT together with either PTEN siRNA or mm siRNA (mm RNA) as designated. Twenty-one hours thereafter, cells were treated with rapamycin (100 nM) for 3 h. The RNA Pol I promoter assays were conducted as described in Materials and Methods. (B) Expression of constitutively activated S6K1 alleviates PTEN-mediated repression of RNA Pol I-dependent transcription. U87 cells were transiently transfected with PrHuCAT (left panel) or −4500/+66 hTBP-luc (right panel), together with either PTEN expression vectors, empty vector, or plasmids expressing constitutively active S6K1 as designated. RNA Pol I and TBP promoter assays were conducted as described for Fig. 1. An example of an autoradiograph is shown above the graphs. The values shown are mean ± SEM for three independent determinations.