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Cloning and characterization of androgen receptor coactivator, ARA55, in human prostate
1George Whipple Lab for Cancer Research, Departments of Pathology, Urology, and Radiation Oncology, University of Rochester Medical Center, Rochester, New York 14642, USA.
Abstract:
Androgen receptor (AR) is a hormone-activated transcriptional factor that can bind to androgen response elements and that regulates the transcription of target genes via a mechanism that presumably involves cofactors. We report here the cloning of a novel AR coactivator ARA55 using a yeast two-hybrid system. ARA55 consists of 444 amino acids with the predicted molecular mass of 55 kDa and its sequence shows very high homology to mouse hic5, a TGF-beta1-inducible gene. Yeast and mammalian two-hybrid systems and co-immunoprecipitation assays all prove ARA55 can bind to AR in a ligand-dependent manner. Transient transfection assay in prostate cancer DU145 cells further demonstrates that ARA55 can enhance AR transcriptional activity in the presence of 1 nM dihydrotestosterone or its antagonists such as 100 nM 17beta-estradiol or 1 microM hydroxyflutamide. Our data also suggest the C-terminal half of ARA55, which includes three LIM motifs, is sufficient to interact with AR. Northern blot and polymerase chain reaction quantitation showed ARA55 can be expressed differently in normal prostate and prostate tumor cells. Together, our data suggests that ARA55 may play very important roles in the progression of prostate cancer by the modulation of AR transactivation.
Insights
Researchers identified a new protein, ARA55, that interacts with the androgen receptor (AR). This interaction enhances AR
Area of Science:
- Molecular Biology
- Endocrinology
- Cancer Research
Background:
- The androgen receptor (AR) is a key regulator of gene transcription, influenced by ligand binding and cofactor interactions.
- Understanding AR cofactors is crucial for deciphering AR-mediated gene regulation, particularly in prostate cancer.
Purpose of the Study:
- To identify and characterize novel coactivators of the androgen receptor.
- To investigate the role of a newly identified coactivator, ARA55, in AR transcriptional activity and prostate cancer.
Main Methods:
- Yeast two-hybrid screening to identify AR-interacting proteins.
- Mammalian two-hybrid assays and co-immunoprecipitation to confirm AR-ARA55 binding.
- Transient transfection assays in prostate cancer cells to assess AR transcriptional activity.
- Northern blot and quantitative PCR to analyze ARA55 expression in prostate tissues.
Main Results:
- A novel AR coactivator, ARA55 (444 amino acids, 55 kDa), was cloned and found to be homologous to mouse hic5.
- ARA55 binds to the AR in a ligand-dependent manner, confirmed by yeast/mammalian two-hybrid and co-immunoprecipitation.
- ARA55 enhances AR transcriptional activity in prostate cancer cells, even in the presence of AR antagonists.
- The C-terminal half of ARA55, containing three LIM motifs, is sufficient for AR interaction.
- Differential expression of ARA55 was observed between normal prostate and prostate tumor cells.
Conclusions:
- ARA55 is a novel ligand-dependent AR coactivator.
- ARA55 enhances AR transactivation, suggesting a role in modulating AR activity.
- Differential expression implies ARA55 may contribute to prostate cancer progression through AR pathway modulation.