The p53-induced Gene-6 (Proline Oxidase) Mediates Apoptosis through a Calcineurin-dependent Pathway

Abstract

Proline oxidase is a p53-induced redox gene that can generate reactive oxygen species (ROS) and mediate apoptosis in tumor cells. We report that proline oxidase is a downstream effector in p53-mediated activation of the calcium/calmodulin-dependent phosphatase calcineurin in lung, renal, colon, and ovarian carcinoma cells. The activation of calcineurin by p53 and proline oxidase was detected by activation of the nuclear factor of activated T cells (NFAT), an established indicator of activated calcineurin. Both proline oxidase-and p53-induced activation of NFAT were sensitive to the calcineurin inhibitors cyclosporin A and FK-506, to scavengers of ROS, and to inhibitors of calcium mobilization. A proline oxidase antisense vector suppressed the ability of p53 to up-regulate proline oxidase, activate calcineurin, and induce apoptosis. Moreover, two renal carcinoma-derived mutant p53 proteins were deficient in inducing proline oxidase expression and in activating calcineurin. Inhibitors of calcineurin and calcium mobilization abolished proline oxidase-mediated apoptosis and reduced p53-induced apoptosis. Treatment of colon and ovarian carcinoma cells with the anticancer genotoxic agent etoposide upregulated both p53 and proline oxidase, activated calcineurin, and induced apoptosis. The etoposide-mediated activation of calcineurin and induction of apoptosis was markedly suppressed by FK-506 calcineurin inhibitor. We propose that proline oxidase mediates apoptosis through the generation of proline-dependent ROS, which then mobilize calcium and activate calcineurin. The activation of calcineurin-regulated transcription factor pathways by proline oxidase might affect gene expression events important to p53 regulation of cell growth and apoptosis.

Proline modulates the intracellular redox environment and protects mammalian cells against oxidative stress

“…Proline metabolism has generally been considered to generate a more oxidative environment via mitochondrial PRODH activity [3][4][5][6]47,63,64]. Here, we have shown that PRODH generates superoxide directly during turnover with proline.…”

“…Of the proline metabolic genes used in this study, most is known about the regulation of PRODH gene expression in cellular apoptosis. Different studies have established that PRODH is activated by p53 and the peroxisome proliferator-activated receptor gamma (PPARγ) regulator with increased PRODH expression leading to oxidative stress [3][4][5][6][7][8]. Hepatocyte nuclear factors HNF4α and HNF1α have also been found to regulate proline degradation [61].…”

“…P5C is then converted to glutamate in a NAD + -dependent reaction catalyzed by P5CDH [2]. Phang and others have shown that in mammalian cells PRODH expression is activated by p53 with proline oxidation leading to increases in intracellular reactive oxygen species (ROS) and induction of cellular apoptosis in various cancer cell lines [3][4][5][6][7][8][9]. Thus, proline serves as a prooxidant in apoptotic signaling via PRODH activity which may be an important mechanism for reducing carcinogenesis [6].…”

Identification of differentially expressed genes in clear cell renal cell carcinoma by analysis of full-length enriched cDNA library

“…It has been reported that metabolism genes, for example proline oxidase [28,29] and cytosolic phospholipase A2 [30,31], were differentially expressed in tumor cells and might have a role in carcinogenesis. Many metabolism genes with abnormal expression in ccRCC tissues, including ASS, were identified by our investigation.…”

“…The oligo-capping method was performed as described previously [29]. Briefly, the phosphate of the 5¢-ends of truncated mRNAs was hydrolyzed by bacterial alkaline phosphatase (Takara).…”