Increased cytochrome P450 17alpha-hydroxylase promoter function in theca cells isolated from patients with polycystic ovary syndrome involves nuclear factor-1.
Cytochrome P450 17alpha-hydroxylase (CYP17) gene expression and androgen biosynthesis are persistently elevated in theca cells isolated from ovaries of women with polycystic ovary syndrome (PCOS). We previously reported that -235 to -109 bp of the CYP17 promoter confers increased CYP17 promoter function in PCOS theca cells. In this report, additional deletion and mutational analyses of the CYP17 promoter were performed to identify the sequences that contribute to increased CYP17 promoter function in PCOS theca cells. Results of these analyses established that augmented promoter function in PCOS theca cells results from preferentially increased basal regulation conferred by sequences between -188 and -147 bp of the CYP17 promoter. Scanning mutant analysis demonstrated that mutations within a 16-bp sequence, spanning -174 to -158 bp of the promoter, ablated increased basal CYP17 promoter function in PCOS theca cells. EMSA analysis demonstrated that the NF-1 family member, NF-1C, bound this sequence. Cotransfection of several NF-1C isoforms expressed in normal and PCOS cells repressed CYP17 promoter function. NF-1C protein and DNA binding were reduced in PCOS theca cell nuclear extracts, as compared with normal. Another NF-1C site between -102 and -90 bp of the promoter was also identified. However, mutation of this site had no effect on differential promoter function in PCOS theca cells. These studies demonstrate that 1) augmented CYP17 promoter function in PCOS theca cells results from increased basal regulation, and 2) diminished NF-1C-dependent repression may be one mechanism underlying increased basal CYP17 promoter activity and altered gene expression in PCOS theca cells.
Wickenheisser
Jessica K
JK
Department of Cellular and Molecular Physiology, Pennsylvania State University College of Medicine, 500 University Drive H166, Hershey, Pennsylvania 17033, USA.
Nelson-DeGrave
Velen L
VL
Quinn
Patrick G
PG
McAllister
Jan M
JM
D000328:Adult; D001483:Base Sequence; D001665:Binding Sites; D022762:CCAAT-Enhancer-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D002478:Cells, Cultured; D005260:Female; D005786:Gene Expression Regulation; D006801:Humans; D008969:Molecular Sequence Data; D009154:Mutation; D051839:NFI Transcription Factors; D011085:Polycystic Ovary Syndrome / Q000235:genetics*; D011401:Promoter Regions, Genetic; D012016:Reference Values; D017384:Sequence Deletion; D013254:Steroid 17-alpha-Hydroxylase / Q000235:genetics* / Q000378:metabolism; D013799:Theca Cells / Q000473:pathology / Q000502:physiology*; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*
D022762:CCAAT-Enhancer-Binding Proteins; D051839:NFI Transcription Factors; D014157:Transcription Factors; D013254:Steroid 17-alpha-Hydroxylase