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A novel androgen receptor mutant, A748T, exhibits hormone concentration-dependent defects in nuclear accumulation and
Alaina J James1, Irina U Agoulnik, Jonathan M Harris
1Department of Molecular and Cellular Biology, Baylor College of Medicine, Houston, TX 77030, USA.
Abstract:
Functional analysis of androgen receptor (AR) gene mutations isolated from prostate cancer has led to the identification of residues that play important roles in the structure and function of the receptor. Here we report the characteristics of a novel AR mutation A748T located in helix 5 of the ligand-binding domain, which was identified in metastatic prostate cancer. Despite a normal hormone-binding affinity, A748T causes hormone concentration-dependent defects in nuclear accumulation and transcriptional activation. Moreover, when equivalent amounts of DNA are transfected, the mutant is expressed at much lower levels than the wild-type AR (ARWT). Treatment with geldanamycin to disrupt receptor-heat shock protein complexes rapidly decreases the levels of ARWT but not A748T, suggesting that the lower expression and rapid degradation rate of A748T is due to weaker interactions with heat shock proteins. Further analysis revealed that hormone dissociates from A748T five times faster than from ARWT. Loss of the ability to form stable amino/carboxyl-terminal interactions causes accelerated dissociation rates in some AR mutants. However, A748T exhibits normal amino/carboxyl-terminal interactions at high hormone concentrations, suggesting that the mutation alters interactions with ligand. Consistent with this conclusion, our structural model predicts that A748T disrupts crucial contact points with ligand, thereby altering the conformation of the ligand-binding domain.
Insights
A novel androgen receptor (AR) mutation, A748T, found in prostate cancer, impairs receptor function and stability. This mutation affects nuclear accumulation and gene activation, despite normal hormone binding.
Area of Science:
- Molecular Biology
- Genetics
- Oncology
Background:
- Androgen receptor (AR) gene mutations are critical in prostate cancer progression.
- Understanding AR mutations aids in developing targeted therapies.
Purpose of the Study:
- To characterize a novel AR mutation, A748T, identified in metastatic prostate cancer.
- To investigate the functional impact of A748T on AR structure and activity.
Main Methods:
- Site-directed mutagenesis to create A748T AR.
- Transfection studies to assess nuclear accumulation and transcriptional activation.
- Hormone-binding assays and geldanamycin treatment to evaluate receptor stability and interactions.
Main Results:
- A748T mutation causes hormone concentration-dependent defects in nuclear accumulation and transcriptional activation.
- The mutant AR (A748T) exhibits lower expression levels and faster degradation than wild-type AR (ARWT).
- Hormone dissociates five times faster from A748T compared to ARWT, suggesting altered ligand interactions.
Conclusions:
- The A748T mutation disrupts AR function by altering ligand-binding domain conformation and interactions.
- This mutation impacts AR stability and activity, contributing to prostate cancer progression.
- A748T represents a potential therapeutic target in prostate cancer treatment.