ISOFORM-SPECIFIC AMINO-TERMINAL DOMAINS DICTATE DNA-BINDING PROPERTIES OF ROR-ALPHA, A NOVEL FAMILY OF ORPHAN HORMONE NUCLEAR RECEPTORS

Three isoforms of a novel member of the steroid hormone nuclear recept or superfamily related to the retinoic acid receptors have been identi fied. The three isoforms, referred to as ROR alpha 1, ROR alpha 2, and ROR alpha 3, share common DNA- and putative ligand-binding domains bu t are characterized by distinct amino-terminal domains generated by al ternative RNA processing. An exon encoding a functionally important su bregion of the amino-terminal domain of the ROR alpha 2 isoform reside s on the opposite strand of a cytochrome c-processed pseudogene. Bindi ng site selection using in vitro-synthesized proteins reveals that the ROR alpha 1 and ROR alpha 2 isoforms bind DNA as monomers to hormone response elements composed of a 6-bp AT-rich sequence preceding a half -site core motif PuGGTCA (RORE). However, ROR alpha 1 and ROR alpha 2 display different binding specificities: ROR alpha 1 binds to and cons titutively activates transcription from a large subset of ROREs, where as ROR alpha 2 recognizes ROREs with strict specificity and displays w eaker transcriptional activity. The differential DNA-binding activity of each isoform maps to their respective amino-terminal domains. Where as truncation of the amino-terminal domain diminishes the ability of R OR alpha 1 to bind DNA, a similar deletion relaxes ROR alpha 2-binding specificity to that displayed by ROR alpha 1. Remarkably, transfer of the entire amino-terminal region of ROR alpha 1 or amino-terminal del etion of ROR alpha 2 confers RORE-binding specificities to heterologou s receptors. These results demonstrate that the amino-terminal domain and the zinc finger region work in concert to confer high affinity and specific DNA-binding properties to the ROR isoforms and suggest a nov el strategy to control DNA-binding activity of nuclear receptors.