Proline oxidase silencing inhibits p53-dependent apoptosis in MCF-7 breast cancer cells

Abstract

Proline oxidase (POX) is mitochondrial proline-degrading enzyme of dual apoptosis/survival function. POX expression and proline availability are considered an underlying mechanism for differential POX functions. The mechanism for POX-dependent regulation of cell death/survival was studied in wild-type (MCF-7^WT) and shRNA POX-silenced breast cancer cells (MCF-7^iPOX). Proline concentration and proteomic analyses were determined by LC/MS/QTOF and LC/MS/ORBITRA, respectively. Inhibition of collagen biosynthesis (proline utilizing process) by 2-methoxyestradiol (2ME) contributed to induction of apoptosis in MCF-7^WT cells, as detected by increase in the expression of active caspase-3, -9 and p53. The process was not shown in MCF-7^iPOX. In MCF-7^iPOX cells prolidase activity and expression as well as proline concentration were drastically increased, compared to MCF-7^WT cells. Down-regulation of p53 in MCF-7^iPOX cells was corroborated by proteomic analysis showing decrease in the expression of p53-related proteins. The mechanism for down-regulation of p53 expression in MCF-7^iPOX cells was found at the level of p53-PEPD complex formation that was counteracted by hydrogen peroxide treatment. In this study, we found that silencing POX modulate pro-survival phenotype of MCF-7 cells and suggest that the mechanism of this process undergoes through down-regulation of p53-dependent signaling.

Keywords: Apoptosis; MCF-7 breast cancer cells; Proline; Proline dehydrogenase; Proline oxidase; p53.