Protein Phosphatases Modulate p53-Bak Pathway to Promote Survival of Human Lens Epithelial Cells

Abstract

Purpose: : Our recent study has shown that through dephosphorylation, protein serine/threonine phosphatase-2A negatively regulate p53 and its downstream proapoptotic gene, bak in mouse to attenuate apoptosis (Qin et al., 2008. Cancer Res. 68:4150-4162). Whether the protein serine/threonine phosphatases regulate p53-Bak pathway in human lens epithelial cells remains to be established.

Methods: : Bioinformatics was used to analyze the human bak promoter and two putative p53 binding sites were identified. Gel mobility shifting assay was used to demonstrate that p53 directly binds to the oligo containing the p53 binding site from the bak promoter. In vitro mutagenesis and dose-dependent overexpression of exogenous p53 were used to confirm that p53 directly regulates Bak in human lens epithelial cells.

Results: : PP-1 and/or PP-2A can directly dephosphorylate p53 at multiple residues Dephosphorylation of p53 at these sites distinctly modulates its ability for DNA binding to the downstream target gene, bak and others, and thus changes its control over Bak and a panel of other downstream genes including p21, PCNA, Bcl-2, and Bax, which are involved in regulation of lens differentiation and apoptosis both in vitro and in vivo.

Conclusions: : Through dephosphorylation of p53, PP-1 and PP-2A directly modulate p53-Bak pathway to promote survival of human lens epithelail cells. One of the major mechanisms for PP-1 to promote survival and thus to maintain transparency of the ocular lens is to negatively regulate the function of p53, which plays a key role in controlling both lens development and cataractogenesis.

Keywords: cell survival • apoptosis/cell death • gene/expression