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Published on: June 27, 2020
FOXA1 modulates EAF2 regulation of AR transcriptional activity, cell proliferation, and migration in prostate cancer
Wenhuan Guo1, Anne L Keener, Yifeng Jing
1Pathology Center, Shanghai First People's Hospital / Faculty of Basic Medicine, Shanghai Jiao Tong University School of Medicine, Shanghai, China; Department of Urology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania.
Background:
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 real-time RT-PCR and luciferase reporter assays. The effect of EAF2 and/or FOXA1 knockdown on LNCaP human prostate cancer cell proliferation and migration was tested using BrdU 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 level, whereas transfected GFP-EAF2 down-regulated the FOXA1 protein. Also, EAF2 knockdown enhanced the expression of AR-target genes, cell proliferation, and migration in LNCaP cells. However, FOXA1 knockdown inhibited the effect of EAF2 knockdown on AR-target gene expression, cell proliferation, and migration in LNCaP 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.
Insights
ELL-associated factor 2 (EAF2) acts as a tumor suppressor in the prostate. This study reveals EAF2 interacts with FOXA1, influencing androgen receptor (AR) signaling, cell proliferation, and migration, key mechanisms in suppressing prostate cancer.
Area of Science:
- Oncology
- Molecular Biology
- Genetics
Background:
- ELL-associated factor 2 (EAF2) is an androgen-regulated tumor suppressor in the prostate.
- The precise mechanisms of EAF2's tumor suppressive function remain largely unelucidated.
- Identifying modulators of EAF2 function is crucial for understanding its role in cancer.
Purpose of the Study:
- To identify factors that modulate EAF2 function.
- To investigate the interaction between EAF2 and FOXA1 in prostate cancer.
- To elucidate the role of EAF2 and FOXA1 in regulating androgen receptor (AR) signaling, cell proliferation, and migration.
Main Methods:
- Utilized a C. elegans model (eaf-1 mutant) and RNAi screening to identify synergistic factors.
- Employed co-immunoprecipitation and protein stability assays to study EAF2-FOXA1 interaction and protein levels in human cells.
- Assessed the impact of EAF2 and FOXA1 modulation on AR-target gene expression, LNCaP cell proliferation, and migration using RT-PCR, luciferase assays, BrdU, and transwell assays.
Main Results:
- Identified pha-4 (C. elegans FOXA1 ortholog) as a factor whose knockdown synergistically enhances the fertility defect of eaf-1 mutants.
- Confirmed EAF2 co-immunoprecipitates with FOXA1, and EAF2 modulates FOXA1 protein levels.
- Demonstrated that EAF2 knockdown enhances AR-target gene expression, cell proliferation, and migration, effects partially dependent on FOXA1.
Conclusions:
- FOXA1 plays a critical role in mediating EAF2's tumor-suppressive effects in prostate cancer.
- Regulation of the AR signaling pathway, cell proliferation, and migration by FOXA1 is a key mechanism for EAF2's suppression of prostate carcinogenesis.
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