Prostate cancer (PC) is the most commonly diagnosed cancer and is the second leading cause of cancer deaths among men in the United States. The ability of tumor cells to survive in the low oxygen (hypoxic) environment created by the expansion of primary tumors is fundamental to their growth in situ as well as metastatic progression that underlies the majority of disease‐related mortality. Prostate Specific Membrane Antigen (PSMA), a type II transmembrane peptidase, is progressively upregulated in ~80% of tumors during PC progression where it correlates negatively with prognosis. We have previously reported that increased PSMA expression on tumor cells alters signal transduction mechanisms downstream of growth factor receptors to promote tumor progression. In addition, we found that expression of PSMA promotes increased survival in hypoxic environments and the current study was designed to determine these PSMA‐dependent survival mechanisms. Using the CRISPR/Cas9 system, we engineered a panel of human and murine PC cell lines (LnCaP, 22rv1, TRAMP‐C1) to lack PSMA expression. Interestingly, loss of PSMA (PSMAKO) in these highly metastatic PC cell lines resulted in a significantly lower proliferation rate than PSMA expressing cells (PSMAWT), formation of smaller anchorage‐independent colonies in soft agar, a shift in the balance of RIPK3/MLKL family of necrosis proteins toward a decreased survival, and a decrease in the migration and intravasation/extravasation potential. Furthermore, in a NOD/SCID xenograft mouse model, LnCaP‐PSMAWT cells formed large, well‐vascularized tumors whereas LnCaP‐PSMAKO showed no evidence of tumor growth, indicating that PSMA is a potent tumor promoter both in vitro and in vivo. Mimicking hypoxic conditions in vitro by treating both PSMAWT and PSMAKO cells with a HiF1 inhibitor, (CoCl2), produced drastic differences in the hypoxia‐induced regulation of the calcineurin A and calpain‐2 calcium‐associated signal‐transduction pathways as well as actin regulated cytoskeletal rearrangement highlighting PSMAs role in resistance to hypoxia as well as its potential contribution to a pro‐tumor phenotype and increased metastatic potential. In support of this notion, in silico meta‐analysis of publicly available human gene expression data sets indicated that changes in PSMA and calcium associated genes correlates significantly with PC metastasis to bone and lymph. Dramatic changes in calcium levels are a primary response of many cells to hypoxia and typically stimulate cell death. These results suggest that PSMA expression on tumor cells confers resistance to hypoxia via changes in the calcium signal‐transduction cascade, thus making the complete disruption of PSMA a promising therapeutic option in management and treatment of PC.Support or Funding InformationResearch reported in this paper was supported by the National Institute of General Medical Sciences of the National Institutes of Health under linked Award Numbers RL5GM118969, TL4GM118971, and UL1GM118970. Support was also received from the National Cancer Institute of the National Institutes of Health K01 Mentored Research Scientist Award to Promote Diversity under linked Award Number K01CA188412. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.