FOXA1 Modulates EAF2 Regulation Of AR Transcriptional Activity,Cell Proliferation,And Migration In Prostate Cancer Cells

Objectives: ELL-associated factor 2(EAF2) is an androgen-regulated tumor suppressor in the prostate. However, the mechanisms underlying tumor suppressive function of EAF2 are still largely unknown. Identification of factors capable of modulating EAF2 function will help elucidate the mechanisms underlying EAF2 tumor suppressive function.METHODS. Using eaf-1(the ortholog of EAF2) mutant C. elegans model, RNAi screen was used to identify factors on the basis of their knockdown to synergistically enhance the reduced fertility phenotype of the eaf-1 mutant C. elegans. In human cells, the interaction of EAF2 with FOXA1 and the effect of EAF2 on the FOXA1 protein levels were determined by co-immunoprecipitation and protein stability assay. The effect of EAF2 and/or FOXA1 knockdown on the expression of AR-target genes was determined by realtime RT-PCR and luciferase reporter assays. The effect of EAF2 and/or FOXA1 knockdown on the ability of AR binding to ARE was tested using Ch IP assay. The effect of EAF2 and/or FOXA1 knockdown on LNCa P human prostate cancer cell proliferation and migration was tested using Brd U incorporation assay and transwell migration assay.RESULTS. RNAi screen identified pha-4, the C. elegans ortholog of mammalian FOXA1, on the basis of its knockdown to synergistically enhance the reduced fertility phenotype of the eaf-1 mutant C. elegans causing sterility. EAF2 co-immunoprecipitated with FOXA1. EAF2 knockdown enhanced endogenous FOXA1 protein levels, whereas transfected GFP-EAF2 down-regulated the FOXA1 protein. Also, EAF2 knockdown enhanced the expression of AR-target genes, AR binding to ARE, cell proliferation, and migration in LNCa P cells. However, FOXA1 knockdown inhibited the effect of EAF2 knockdown on AR-target gene expression, AR binding to ARE, cell proliferation, and migration in LNCa P cells, suggesting that FOXA1 can modulate EAF2 regulation of AR transcriptional activation, cell proliferation, and migration.CONCLUSIONS. These findings suggest that regulation of the AR signaling pathway, cell proliferation, and migration through FOXA1 represents an important mechanism of EAF2 suppression of prostate carcinogenesis.