Protein phosphatase 1 activation and alternative splicing of Bcl-x and Mcl-1 by EGCG plus ibuprofen

Epigallocatechin-3-gallate (EGCG) and ibuprofen synergistically act to suppress proliferation and enhance apoptosis of prostate cancer cell lines, PC-3 and LNCaP. The purpose of this study was to investigate the mechanism of underlying this synergism. Most interestingly, EGCG+ ibuprofen treatment in PC-3 cells resulted in altering the ratio of the splice variants of Bcl-X and Mcl-1, downregulating the mRNA levels of anti-apoptotic Bcl-X(L) and Mcl-1 (L) with a concomitant increase in the mRNA levels of pro-apoptotic Bcl-X(s) and Mcl-I (s). However, there were no apparent changes in splicing variants in either ibuprofen or EGCG treated cells. Induction of alternative splicing was correlated with increased activity of protein phosphatase I (PP1) in EGCG + ibuprofen-treated cells, since pretreatment with calyculin A and tautomycin blocked EGCG + ibuprofen-incluced alternative splicing in PC-3 cells in contrast to pretreatment with okadaic acid. On the other hand, EGCG + ibuprofen treatment in LNCaP cells did not alter splicing variants of Bcl-X and Mcl-1, despite the increase in protein phosphatase activity. In both cell lines, EGCG+ibuprofen inhibited cell proliferation synergistically. Taken together, this study demonstrate for the first time that EGCG + ibuprofen upregulated PP1 activity, which in turn induced alternative splicing of Bcl-X and Mcl-I in a cell-type specific manner. Our study also demonstrates that the activation of PP1 contributes to the alternative splicingof Mcl-I