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Molecular Endocrinology 11 (2): 218-228
Copyright © 1997 by The Endocrine Society

The N-Terminal Domain of Transcription Factor IIB Is Required for Direct Interaction with the Vitamin D Receptor and Participates in Vitamin D-Mediated Transcription

Hisashi Masuyama, Stephen C. Jefcoat, Jr. and Paul N. MacDonald

St. Louis University Health Sciences Center Department of Pharmacological and Physiological Science St. Louis, Missouri 63104

The interaction of the vitamin D receptor (VDR) with transcription factor IIB (TFIIB) represents a potential physical link between the VDR-DNA complex and the transcription preinitiation complex. However, the functional relevance of the VDR-TFIIB interaction in vitamin D-mediated transcription is not well understood. In the present study, we used site-directed mutagenesis to demonstrate that the structural integrity of the amino-terminal zinc finger of TFIIB is essential for VDR-TFIIB complex formation. Altering the putative zinc-coordinating residues (C15, C34, C37, or H18) to serines abolished TFIIB interaction with the VDR as assessed in a yeast two-hybrid system and by in vitro protein interaction assays. This N-terminal, VDR-interactive domain functioned as a selective, dominant-negative inhibitor of vitamin D-mediated transcription. Expressing amino acids 1–124 of human TFIIB (N-TFIIB) in COS-7 cells or in osteoblastic ROS17/2.8 cells effectively suppressed 1,25-dihydroxyvitamin D3 (1,25-(OH)2D3)-induced transcription, but had no effect on basal or glucocorticoid-activated transcription. A mutant N-terminal domain [N-TFIIB(C34S:C37S)] that does not interact with VDR had no impact on 1,25-(OH)2D3-induced transcription. Interestingly, both in vitro and in vivo protein interaction assays showed that the VDR-TFIIB protein complex was disrupted by the 1,25-(OH)2D3 ligand. Mechanistically, these data establish a functional role for the N terminus of TFIIB in VDR-mediated transcription, and they allude to a role for unliganded VDR in targeting TFIIB to the promoter regions of vitamin D-responsive target genes.




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