The transcriptional regulating protein of 132 kDa (TReP-132) enhances P450scc gene transcription through interaction with steroidogenic factor-1 in human adrenal cells.
The human P450scc gene is regulated by the tissue-specific orphan nuclear receptor, steroidogenic factor-1 (SF-1), which plays a key role in several physiologic processes including steroid synthesis, adrenal and gonadal development, and sexual differentiation. Several studies have demonstrated the interaction of SF-1 with different proteins. However, it is clear that additional factors not yet identified are involved with SF-1 to regulate different target genes. Recently, it was demonstrated that a novel transcriptional regulating protein of 132 kDa (TReP-132) regulates expression of the human P450scc gene. The overexpression of TReP-132 in adrenal cells increases the production of pregnenolone, which is associated with the activation of P450scc gene expression. Considering the colocalization of TReP-132 and SF-1 in steroidogenic tissues such as the adrenal and testis, and the presence of two putative LXXLL motifs in TReP-132 that can potentially interact with SF-1, the relationship between these two factors on the P450scc gene promoter was determined. The coexpression of SF-1 and TReP-132 in adrenal NCI-H295 cells cooperates to increase promoter activity. Pull-down experiments demonstrated the interaction between TReP-132 and SF-1, and this was further confirmed in intact cells by coimmunoprecipitation/Western blot and two-hybrid analyses. Deletions and mutations of the TReP-132 cDNA sequence demonstrate that SF-1 interaction requires the LXXLL motif found at the amino-terminal region of the protein. Also, the "proximal activation domain" and the "AF-2 hexamer" motif of SF-1 are involved in interaction with TReP-132. Consistent with previous studies showing interaction between CBP/p300 and SF-1 or TReP-132, the coexpression of these three proteins results in a synergistic effect on P450scc gene promoter activity. Taken together the results in this study identify a novel function of TReP-132 as a partner in a complex with SF-1 and CBP/p300 to regulate gene transcription involved in steroidogenesis.
Gizard
Florence
F
Oncology and Molecular Endocrinology Research Center, Laval University, Québec GIK 7P4, Canada.
Lavallee
Bernard
B
DeWitte
Frederique
F
Teissier
Elisabeth
E
Staels
Bart
B
Hum
Dean W
DW
D000310:Adrenal Gland Neoplasms / Q000378:metabolism; D000311:Adrenal Glands / Q000166:cytology*; D020816:Amino Acid Motifs; D000818:Animals; D015153:Blotting, Western; D002786:Cholesterol Side-Chain Cleavage Enzyme / Q000235:genetics / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D018076:DNA, Complementary / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism* / Q000502:physiology*; D050881:E1A-Associated p300 Protein; D051717:Fushi Tarazu Transcription Factors; D005982:Glutathione Transferase / Q000378:metabolism; D018398:Homeodomain Proteins; D006801:Humans; D008156:Luciferases / Q000378:metabolism; D051379:Mice; D008957:Models, Genetic; D009687:Nuclear Proteins / Q000378:metabolism; D010957:Plasmids / Q000378:metabolism; D011233:Precipitin Tests; D011284:Pregnenolone / Q000378:metabolism; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D018160:Receptors, Cytoplasmic and Nuclear; D054339:Steroidogenic Factor 1; D013997:Time Factors; D015534:Trans-Activators / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism* / Q000502:physiology*; D014158:Transcription, Genetic; D014162:Transfection; D014407:Tumor Cells, Cultured
D018076:DNA, Complementary; D004268:DNA-Binding Proteins; D051717:Fushi Tarazu Transcription Factors; D018398:Homeodomain Proteins; C515057:NR5A1 protein, human; D009687:Nuclear Proteins; D018160:Receptors, Cytoplasmic and Nuclear; D054339:Steroidogenic Factor 1; C436322:TRERF1 protein, human; D015534:Trans-Activators; D014157:Transcription Factors; C486668:Trerf1 protein, mouse; C515058:steroidogenic factor 1, mouse; D011284:Pregnenolone; D008156:Luciferases; D002786:Cholesterol Side-Chain Cleavage Enzyme; D050881:E1A-Associated p300 Protein; C494361:Ep300 protein, mouse; D005982:Glutathione Transferase