Transition from monomeric to homodimeric DNA binding by nuclear receptors: identification of RevErbAalpha determinants required for RORalpha homodimer complex formation.

Abstract

Nuclear hormone receptors belong to a class of transcription factors that recognize specific DNA sequences either as monomers, homodimers, or heterodimers with the common partner retinoic X receptor. In vitro mutagenesis studies, as well as determination of the crystal structure of several complexes formed by the DNA-binding domain of receptors bound to their cognate response elements, have begun to explain the molecular basis for protein-DNA and protein-protein interactions essential for high-affinity and specific DNA binding by nuclear receptors. In this study, we have used the related orphan nuclear receptors, RORalpha and RevErbAalpha, to study the molecular determinants involved in the transition from monomeric to homodimeric modes of DNA binding by nuclear receptors. While both receptors bind DNA as monomers to a response element containing a core AGGTCA half-site preceded by a 5'-A/T-rich flanking sequence, RevErbAalpha also binds as a homodimer to an extended DR2 element. Gain-of-function experiments using point mutations and subdomain swaps between RORalpha and RevErbAalpha identify four amino acids within RevErbAalpha sufficient to confer RORalpha with the ability to form cooperative homodimer complexes on an extended DR2. This study reveals how the transition from monomer to homodimer DNA binding by members of the nuclear receptor superfamily could be achieved from relatively few amino acid substitutions.