INTERACTION BETWEEN THE RETINOID-X-RECEPTOR AND TRANSCRIPTION FACTOR IIB IS LIGAND-DEPENDENT IN-VIVO

The retinoid X receptor (RXR) influences gene activation through heter odimeric and homodimeric association with DNA and associates with TATA binding protein, TAF110, and cAMP response element-binding protein-bi nding protein; yet the molecular mechanisms responsible for gene activ ation by RXRs remain incompletely defined. Since the general transcrip tion factor IIB (TFIIB) is a common target of sequence-specific transc riptional activators, we suspected that RXR might regulate target gene s via an interaction with TFIIB. Coimmunoprecipitation, far Western an alysis, and glutathione S-transferase binding studies indicated that m urine RXR beta (mRXR beta) was capable of binding to human TFIIB in vi tro. Functional analysis with a dual-hybrid yeast system and cotransfe ction assays revealed the interaction of mRXR beta with TFIIB to be li gand-dependent in vivo. Truncation experiments mapped the essential bi nding regions to the carboxyl region of mRXR beta (amino acids (aa) 25 4-389) and two regions in the carboxyl region of TFIIB (aa 178-201 and aa 238-271). Furthermore, the Delta 390-410 mRXR beta mutant bound to TFIIB in vitro but was not active in the dual-hybrid yeast system, su ggesting that the extreme carboxyl region of RXR was required for in v ivo interaction with TFIIB. These data indicate that interaction of mR XR beta with TFIIB is specific, direct, and ligand-dependent in vivo a nd suggest that gene activation by RXR involves TFIIB.