Receptor-interacting protein (RIP) 140 interacts with several nuclear receptors, but its function in regulation of nuclear receptor action has been debated. Here we have examined the role of RIP140 in regulation of Steroidogenic factor-1 (SF-1)-dependent transcription. SF-1 interacts with RIP140 through its activation function-2 (AF-2) domain. Several domains of RIP140 interact directly with SF-1, but the carboxyl-terminal region containing 4 of its 9 LXXLL motifs showed the strongest SF-1 interaction. Coexpression of RIP140 and SF-1 in different cell types demonstrated that RIP140 acts as a potent corepressor of transcription from the SF-1 responsive cAMP regulatory sequence 2 (CRS2) element of the CYP17 gene and a variety of SF-1 responsive promoter genes. RIP140 also counteracted the stimulatory action of p160/SRC coactivators. The inhibitory effect of RIP140 was partially reversed by Trichostatin A, suggesting a role of histone deacetylase (HDAC) activity in RIP140-mediated repression of SF-1. Quantitation of endogenous coregulator mRNA levels revealed cell type specific differences that could affect the repressor action by overexpressed RIP140.