MDM2 regulates the stability of AR, AR-V7, and TM4SF3 proteins in prostate cancer

Androgen receptor (AR) and its constitutively active splice variant, AR Variant 7 (AR-V7), regulate genes essential for the development and progression of prostate cancer. Degradation of AR and AR-V7 by the ubiquitination proteasomal pathway is important for the regulation of both their protein stability. Our published results demonstrate that the interaction of TM4SF3 with either AR or AR-V7 leads to mutual stabilization due to a reduction in their ubiquitination and proteasomal degradation. These results led us to search for a common E3 ligase for AR, AR-V7, and TM4SF3. Depletion by siRNA of several E3 ligases identified MDM2 as the common E3 ligase. MDM2 inhibition by siRNA depletion or using a pharmacological inhibitor (MDM2i) of its E3 ligase activity led to elevated levels of endogenous AR, AR-V7, and TM4SF3 in prostate cancer cells. MDM2 knockdown in PC-3 cells, which do not express AR, also increased TM4SF3, demonstrating that MDM2 affects the TM4SF3 protein independent of AR. We further demonstrate that MDM2i treatment reduced the ubiquitination of AR and TM4SF3, suggesting that MDM2 can induce the ubiquitination of these proteins. Increased AR and AR-V7 protein levels induced by MDM2i treatment resulted in the expected increased expression of AR-regulated genes and enhanced proliferation and migration of both LNCaP and Enzalutamide-resistant CWR-22Rv1 prostate cancer cells. Thus, our study expands the known roles of MDM2 in prostate cancer to include its potential involvement in the important mutual stabilization that TM4SF3 exhibits when interacting with either AR or AR-V7.