Research Paper Volume 16, Issue 5 pp 4116—4137

Figure 4. PR55α expression increases RB phosphorylation in normal and malignant pancreatic cells. (A) A schematic depiction of the regulation of the p16/RB cascade by PR55α-controlled PP2A enzymes. PR55α suppresses the expression of p16, which acts as the primary inhibitor of the CDK4/6 kinases that phosphorylate and inactivate RB, thereby resulting in its dissociation from the E2F transcriptional factor. The net effect of PR55α is promoting the G1/S transition by allowing the phosphorylation of RB and the release of E2F. (B) HPNE cells transduced with Dox-inducible PR55α, or control vector were incubated with 1 μg/ml Dox for 3 days to induce ectopic PR55α expression. The cells were exposed to 10 Gy IR and then incubated for the times indicated. Harvested cells were analyzed for RB-Ser708 phosphorylation, total RB level, and GAPDH (as an internal control). (C) CD18/HPAF-transduced with the Control or PR55α shRNA were incubated with 2 μg/ml Dox for 5 days to knockdown PR55α expression, after which cells were analyzed by immunoblotting for differences in RB-Ser708 phosphorylation, total RB, and GAPDH.