CHARACTERIZATION OF RECEPTOR INTERACTION AND TRANSCRIPTIONAL REPRESSION BY THE COREPRESSOR SMRT

SMRT (silencing mediator of retinoic acid and thyroid hormone receptor ) and N-CoR (nuclear receptor corepressor) are two related transcripti onal corepressors that contain separable domains capable of interactin g with unliganded nuclear receptors and repressing basal transcription . To decipher the mechanisms of receptor interaction and transcription al repression by SMRT/N-CoR, we have characterized protein-protein int eracting surfaces between SMRT and nuclear receptors and defined trans criptional repression domains of both SMRT and N-CoR. Deletional analy sis reveals two individual nuclear receptor domains necessary for stab le association with SMRT and a C-terminal helix essential for corepres sor dissociation. Coordinately, two SMRT domains are found to interact independently with the receptors. Functional analysis reveals that SM RT contains two distinct repression domains, and the corresponding reg ions in N-CoR also repress basal transcription. Both repression domain s in SMRT and N-CoR interact weakly with mSin3A, which in turn associa tes with a histone deacetylase HDAC1 in a mammalian two-hybrid assay. Far-Western analysis demonstrates a direct protein-protein interaction between two N-CoR repression domains with mSin3A. Finally we demonstr ate that overexpression of full-length SMRT further represses basal tr anscription from natural promoters. Together, these results support a role of SMRT/N-CoR in corepression through the utilization of multiple mechanisms for receptor interactions and transcriptional repression.