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Mitochondrial localization, import, and mitochondrial function of the androgen receptor.

ABSTRACT: Nuclear localization of androgen receptor (AR) directs transcriptional regulation of a host of genes, referred to as genomic signaling. Additionally, nonnuclear or nongenomic activities of the AR have long been described, but understanding of these activities remains elusive. Here, we report that AR is imported into and localizes to mitochondria and has a novel role in regulating multiple mitochondrial processes. Employing complementary experimental approaches of AR knockdown in AR-expressing cells and ectopic AR expression in AR-deficient cells, we demonstrate an inverse relationship between AR expression and mitochondrial DNA (mtDNA) content and transcription factor A, mitochondrial (TFAM), a regulator of mtDNA content. We show that AR localizes to mitochondria in prostate tissues and cell lines and is imported into mitochondria in vitro We also found that AR contains a 36-amino-acid-long mitochondrial localization sequence (MLS) capable of targeting a passenger protein (GFP) to the mitochondria and that deletion of the MLS abolishes the import of AR into the mitochondria. Ectopic AR expression reduced the expression of oxidative phosphorylation (OXPHOS) subunits. Interestingly, AR also controlled translation of mtDNA-encoded genes by regulating expression of multiple nuclear DNA-encoded mitochondrial ribosomal proteins. Consistent with these observations, OXPHOS supercomplexes were destabilized, and OXPHOS enzymatic activities were reduced in AR-expressing cells and restored upon AR knockdown. Moreover, mitochondrial impairment induced AR expression and increased its translocation into mitochondria. We conclude that AR localizes to mitochondria, where it controls multiple mitochondrial functions and mitonuclear communication. Our studies also suggest that mitochondria are novel players in nongenomic activities of AR.

SUBMITTER: Bajpai P

PROVIDER: S-EPMC6484137 | biostudies-literature | 2019 Apr

REPOSITORIES: biostudies-literature

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Project description:BackgroundCastration-resistant prostate cancer can develop resistance to enzalutamide because of androgen receptor (AR) point mutations, AR overexpression, constitutively active AR splice variants, and/or elevated intratumoral androgen synthesis. The point mutation ARF876L was reported to be stimulated, instead of inhibited, by enzalutamide, thus contributing to enzalutamide resistance. We have recently developed JJ-450 as a novel AR antagonist with the potential to treat enzalutamide-resistant castration-resistant prostate cancer (CRPC).MethodsWe employed several assays to determine the impact of JJ-450 and enzalutamide on prostate cancer cell lines expressing green fluorescent protein (GFP)-ARF876L . These assays include a prostate-specific antigen enhancer/promoter-based luciferase assay to determine AR transcriptional activity, a quantitative real-time polymerase chain reaction assay, and Western blot analysis to detect expression of AR-target genes at the messenger RNA and protein level, fluorescence microscopy to show AR subcellular localization, and a 5-bromo-2'-deoxyuridine assay to measure prostate cancer cell proliferation.ResultsAs expected, enzalutamide inhibited wild-type (WT) AR but not ARF876L transcriptional activity in the luciferase assay. In contrast, JJ-450 inhibited both WT-AR and ARF876L transcriptional activity to a similar extent. Also, enzalutamide retarded androgen-induced nuclear import of GFP-AR, but not GFP-ARF876L , whereas JJ-450 retarded nuclear import of both GFP-AR and GFP-ARF876L . To further evaluate JJ-450 inhibition of ARF876L , we stably transfected C4-2 cells separately with GFP-AR or GFP-ARF876L . Enzalutamide inhibited endogenous AR-target gene expression in C4-2-GFP-ARWT , but not in the C4-2-GFP-ARF876L subline, whereas JJ-450 inhibited AR-target gene expression in both C4-2 sublines. More importantly, enzalutamide inhibited proliferation of C4-2-GFP-ARWT , but not of the C4-2-GFP-ARF876L subline, whereas JJ-450 inhibited proliferation of both C4-2 sublines.ConclusionJJ-450 inhibits enzalutamide-resistant ARF876L mutant nuclear translocation and function. Our findings suggest that JJ-450 and its analogs should be further developed to provide a potential new approach for the treatment of enzalutamide-resistant CRPC.

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Mitochondrial localization, import, and mitochondrial function of the androgen receptor.

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