Thyroid receptor activator molecule, TRAM-1, is an androgen receptor coactivator

An androgen receptor (AR) interacting protein was isolated from a HeLa cell complementary DNA library by two-hybrid screening in yeast using the AR DN A and ligand binding domains [amino acids (aa) 481-919] as bait. AR binding of the protein in yeast was dependent on the presence of testosterone or d ihydrotestosterone (DHT). The isolated protein is identical to thyroid rece ptor activator molecule TRAM-1 but lacking aa 1-458. TRAM-1 is a steroid re ceptor coactivator-3 (SRC-3) subtype. In affinity matrix assays, S-35-label ed TRAM-1 bound the GST-AR ligand binding domain (aa 624-919) and GST-AR N- terminal and DNA binding domains (aa 1-660), but not the GST-AR DNA binding domain (aa 544-634) alone. Coexpression of TRAM-1 increased DHT-dependent AR transactivation 5-fold and constitutive activity of AR (aa 1-660) N-term inal and DNA-binding domains increased g-fold. Full-length TRAM-1 (aa 1-142 4) and the partial (aa 459-1424) were AR and GR coactivators as was SRC-1. In human testis, immunostaining of SRC-3 colocalized with AR in nuclei of S ertoli cells and peritubular myoid cells, indicating it could function as a n AR coactivator in these cells. SRC-3 was also present in nuclei of sperma togenic cells where AR was not expressed, suggesting it might also be a coa ctivator with other nuclear receptors that regulate spermatogenesis.