The Plasma Membrane TRPM8 Plays a Protective Role against Prostate Cancer Progression; Trpm8 Gene as a Downstream Target of P53 Tumor-Suppressor

Abstract

Transient receptor potential melastatin 8 (TRPM8) is a cold-sensing and Ca2+ permeable channel. The mRNA of TRPM8 is overexpressed in early prostate tumors with high androgen levels, while anti-androgen therapy greatly reduces its expression. From the chromatin-immunoprecipitation (ChIP) analysis, we observed that an androgen response element (ARE) mediates androgen regulation of trpm8. Although TRPM8 mRNA is expressed at high levels, we found that the TRPM8 protein undergoes ubiquitination and degradation in prostate cancer (PC) cells. The mass-spectrometry analysis of TRPM8, immunoprecipitated from the membrane and total cell lysates of LNCaP cells identified ubiquitin-like modifier-activating enzyme 1 (UBA1), an enzyme that participates in the initial step of the ubiquitination cascade. We found that PYR-41, a potent inhibitor of UBA1, increased TRPM8 activity on the plasma membrane of LNCaP cells. Furthermore, PYR-41-mediated PMTRPM8 activity was accompanied by enhanced activation of cell cycle inhibitor, p53 and apoptosis associated molecule, Caspase-9. In addition, we found that the promoter region of trpm8 possesses putative binding sites for p53 and that the overexpression of p53 increased the TRPM8 mRNA levels in LNCaP cells. Our findings support previous studies that suggest the balance of androgen receptor (AR) and p53 expression regulates androgen-dependent growth of PC. Alternatively, effect of androgen / AR inhibition / TRPM8 overexpression on cancer cell proliferation and apoptosis was examined by FACS, TUNEL and clonogenic assay. All these experiments indicate that the TRPM8 protein stability is dependent on AR and could be negatively regulated at the post-translational level by AR. TRPM8 overexpression in PC cells showed anti-proliferative effect, suggesting the importance of TRPM8 channel as a tumor regulator.