PKC Alpha Affects Cell Cycle Progression and Proliferation in Human RPE Cells Through the Downregulation of P27kip1

Purpose: Protein kinase C (PKC) plays an important role in the regulation of retinal pigment epithelium (RPE) cell proliferation. In this study, we investigated which of these isozymes could be responsible for the cell cycle and proliferation in human RPE cells.

Methods: The effect of PKC activators on human RPE cell cycle progression was tested by flow cytometry. To identify the isoform of PKC responsible for the increased progression of the cells through the cell cycle, we monitored the effect of phorbol 12-myristate 13-acetate (PMA) on the subcellular localization of the nine PKC isoforms expressed in RPE cells. To evaluate the molecular mechanism by which PKC(alpha) induces cell cycle progression, we examined the transcript, protein, and cellular levels of cell cycle regulatory proteins using RT-PCR, western blotting, and a confocal microscope, respectively.

Results: We demonstrated that PKC activation by PMA affected cell cycle progression in RPE cells. Of the nine PKC isoforms that were present in RPE cells, we found PKC(alpha) was both necessary and sufficient to promote cell cycle progression after being stimulated with PMA. Decreased PKC(alpha) expression resulted in a significant decrease in cell proliferation. The only cell cycle-regulatory molecule whose expression was rapidly altered and decreased by PKC(alpha) activity was the cyclin- dependent kinase (CDK) inhibitor p27(kip1).

Conclusions: These results suggest that PKC(alpha) affects cell cycle progression and proliferation in human RPE cells through the downregulation of p27(kip1).