The promyelocytic leukemia (PML) tumour suppressor protein plays a key role in the cellular stress response. PML is critical for the formation of distinct nuclear structures called PML-nuclear bodies (PML-NBs), important for the tumour suppressive activity of PML. We previously discovered that the E3 ligase E6-associated protein (E6AP) is a major regulator of PML protein stability and PML-NB formation. PML has been extensively studied in human cancers and its expression has been found to be frequently down-regulated or lost. In prostate cancers, the loss of PML is also associated with the development and progression of invasive prostate cancer. Interestingly, the regulation of PML occurs at the protein level, which prompted us to hypothesize that elevated levels of E6AP may lead to the down-regulation of PML, thereby playing a role in the development and progression of human prostate cancer. We have screened prostate cancer samples for the expression of E6AP and PML, and correlated this data with survival following radical prostatectomy. We found that patients with high E6AP and low PML expression levels have the poorest survival rates. To test our hypothesis, we examined the correlation between E6AP and PML expression in a panel of prostate cancer cell lines. We studied the effect of E6AP on PML expression by down-regulation of E6AP using inducible shRNA in these cell lines. The down-regulation of E6AP successfully restored PML expression and enhanced the formation of PML-NBs. Down-regulation of E6AP was also associated with a reduction in cell numbers and an increased sensitization of prostate cancer cells to stress-induced death. Further, using an in vivo xenograft mouse model, we found that down-regulation of E6AP reduced tumour formation. Our study reveals a novel role for the involvement of the E6AP-PML axis in prostate cancer.