Analysis of the Functional Role of Steroid Receptor Coactivator-1 in Ligand-Induced Transactivation by Thyroid Hormone Receptor

M. Jeyakumar
1997 Molecular Endocrinology  
The nuclear hormone receptors belonging to the steroid/thyroid/retinoid receptor superfamily are ligand-inducible transcription factors. These receptors modulate transcription of specific cellular genes, either positively or negatively, by interacting with specific hormone response elements located near the target promoters. Recent studies indicated that the hormone-occupied, DNA-bound receptor acts in concert with a cellular coregulatory factor, termed coactivator, and the basal transcription
more » ... asal transcription machinery to mediate gene activation. Consistent with this scenario, a number of nuclear proteins with potential coactivator function have been isolated. In the present study, we demonstrate that steroid receptor coactivator-1 (SRC-1), a recently isolated candidate coactivator, functions as a positive regulator of the thyroid hormone receptor (TR)-mediated transactivation pathway. In transient transfection experiments, coexpression of SRC-1 significantly enhanced ligand-dependent transactivation of a thyroid hormone response element (TRE)-linked promoter by human TR␤. Our studies revealed that deletion of six amino acids (451-456) in the extreme COOH-terminal region of TR␤ resulted in a receptor that retained the ability to bind T 3 but failed to be stimulated by SRC-1. These six amino acids are part of an amphipathic helix that is highly conserved among nuclear hormone receptors and contains the core domain of the ligand-dependent transactivation function, AF-2. In agreement with this observation, in vitro protein binding studies showed that SRC-1 (451-456) . We demonstrated that a synthetic peptide containing the COOH-terminal amino acids (437-456) of TR␤ efficiently blocked the ligand-induced binding of SRC-1 to the receptor. These results suggest that the conserved amphipathic helix that constitutes the AF-2 core domain of TR␤ is critical for interaction with SRC-1 and thereby plays a central role in coactivator-mediated transactivation. We further observed that a heterodimer of TR␤ and retinoid X receptor-␣ (RXR␣), either in solution or bound to a DR؉4 TRE, recruited SRC-1 in a T 3 -dependent manner. The AF-2 of TR was clearly involved in this process because a TR-RXR heterodimer containing a mutant TR␤ (1-450) with impaired AF-2 failed to bind to SRC-1. Surprisingly, the RXR-specific ligand 9-cis-retinoic acid induced binding of SRC-1 to the RXR component of the TRE-bound heterodimer. This novel finding suggests that RXR, as a heterodimeric partner of TR, has the potential to play an active role in transcriptional regulation. Our results raise the interesting possibility that a RXR-specific ligand may modulate T 3 -mediated signaling by inducing additional interactions between TRE-bound TR-RXR heterodimer and the coactivator. (Molecular Endocrinology 11: 755-767, 1997) Recent studies in several laboratories indicate that nuclear receptors repress or enhance transcription by interacted with a ligand binding domain peptide (145-456) of TR␤ in a T 3 -dependent manner, whereas it failed to interact with a mutant ligand binding domain lacking the amino acids
doi:10.1210/me.11.6.755 pmid:9171239 fatcat:6ub4z44kwnetfpiwhkpnptw3zy