MULTIPLE ORPHAN NUCLEAR RECEPTORS CONVERGE TO REGULATE RAT P450C17 GENE-TRANSCRIPTION - NOVEL MECHANISMS FOR ORPHAN NUCLEAR RECEPTOR ACTION

The orphan nuclear receptor steroidogenic factor-1 (SF-1) plays a key role in regulating the expression of the rat P450c17 gene in testicula r Leydig and in adrenocortical cells. Other DNA sequences, not bound b y SF-l, are also involved in transcriptional regulation of the rat P45 0c17 gene in both cell types. The region from -447/-399 or from -447/- 419 increased both basal and cAMP-induced transcription, and the regio n from -418/ -399 increased basal transcription to a greater extent th an the intact -447/-399 DNA. The -447/-399 DNA sequence contains three imperfect copies of the orphan nuclear receptor-binding motif, AGGTCA , and at least three known orphan nuclear receptors, chicken ovalbumin upstream promoter transcription factor (COUP-TF), SF-1, and an early response gene induced by nerve growth factor (NGFI-B), bind to -447/-3 99 DNA. The AGGTCA triad is bound by one set of nuclear proteins when these three elements are colinear and is bound by a different set of p roteins when these elements are separated. When the elements are separ ated, COUP-TF no longer binds, and the region -418/-399 is bound by a protein that greatly stimulates basal transcription. The region -447/- 419 is bound by two different proteins that mediate both basal and cAM P-stimulated transcription. We call the protein binding to -418/-399 s teroidogenic factor inducer of transcription-1 (StF-IT-1), and one of the proteins binding to -447/-419, StF-IT-2. SF-1 binds to a second AG GTCA element in the -447/-419 region. StF-IT-1 and StF-IT-2 are both f ound in Leydig and adrenal cells, and transcriptional regulation is si milar in both cell types. SF-1 and NGF-IB may increase transcription b y displacing COUP-TF (a transcriptional repressor) because these prote ins share DNA-binding domains. However, neither SF-1 nor NGF-IB alone, binding as monomers, increases transcription. Rather, these proteins must interact with another DNA-binding protein, e.g. StF-IT-2, to incr ease transcription. StF-IT-2 also requires interaction with SF-1 (or N GF-IB) bound to DNA and cannot increase transcription by itself. This mechanism of action is different from the mechanism by which SF-1 regu lates transcription from the -84/-55 region of the rat P450c17 gene. T hus, we have defined a novel mechanism of action for orphan nuclear re ceptors that bind to DNA as monomers.