ISOLATION AND CHARACTERIZATION OF A NOVEL COACTIVATOR PROTEIN, NCOA-62, INVOLVED IN VITAMIN-D-MEDIATED TRANSCRIPTION

The vitamin D receptor (VDR) forms a heterodimeric complex with retino id X receptor (RXR) and binds to vitamin D-responsive promoter element s to regulate the transcription of specific genes or gene networks. Th e precise mechanism of transcriptional regulation by the VDR RXR heter odimer is not well understood, but it may involve interactions of VDR RXR with transcriptional coactivator or corepressor proteins. Here, a yeast two-hybrid strategy was used to isolate proteins that selectivel y interacted with VDR and other nuclear receptors, One cDNA clone desi gnated NCoA-62, encoded a 62,000-Da protein that is highly related to BX42, a Drosophila melanogaster nuclear protein involved in ecdysone-s timulated gene expression. Yeast two-hybrid studies and in vitro prote in-protein interaction assays using glutathione S-transferase fusion p roteins demonstrated that NCoA-62 formed a direct protein-protein cont act with the ligand binding domain of VDR, Coexpression of NCoA-62 in a vitamin D-responsive transient gene expression system augmented 1,25 -dihydroxyvitamin D-3-activated transcription, but it had little or no effect on basal transcription or gal4-VP16-activated transcription. N CoA-62 also interacted with retinoid receptors, and its expression enh anced retinoic acid-, estrogen-, and glucocorticoid-mediated gene expr ession. These data indicate that NCoA-62 may be classified into an eme rging set of transcriptional coactivator proteins that function to fac ilitate vitamin D- and other nuclear receptor-mediated transcriptional pathways.