Alternative splicing of DENND1A, a PCOS candidate gene, generates variant 2.
Polycystic ovary syndrome (PCOS) is a common endocrinopathy characterized by hyperandrogenism and metabolic disorders. The excess androgens may be of both ovarian and adrenal origin. PCOS has a strong genetic component, and genome-wide association studies have identified several candidate genes, notably DENND1A, which encodes connecdenn 1, involved in trafficking of endosomes. DENND1A encodes two principal variants, V1 (1009 amino acids) and V2 (559 amino acids). The androgen-producing ovarian theca cells of PCOS women over-express V2. Knockdown of V2 in these cells reduces androgen production, and overexpression of V2 in normal theca cells confers upon them a PCOS phenotype of increased androgen synthesis. We report that human adrenal NCI-H295A cells express V1 and V2 mRNA and that the V2 isoform is produced by exonization of sequences in intron 20, which generates a unique exon 20A, encoding the C-terminus of V2. As in human theca cells from normal women, forced expression of V2 in NCI-H295A cells resulted in increased abundance of CYP17A1 and CYP11A1 mRNAs. We also found genetic variation in the intronic region 330 bp upstream from exon 20A, which could have the potential to drive the selective expression of V2. There was no clear association with these variants with PCOS when we analyzed genomc DNA from normal women and women with PCOS. Using minigene expression vectors in NCI-H295A cells, this variable region did not consistently favor splicing of the V2 transcript. These findings suggest increased V2 expression in PCOS theca cells is not the result of genomic sequence variation in intron 20.
Department of Pediatrics and the Center for Reproductive Sciences, University of California, San Francisco, United States.
Meng Kian
MK
Tee
Department of Pediatrics and the Center for Reproductive Sciences, University of California, San Francisco, United States.
Mart
M
Speek
Department of Pediatrics and the Center for Reproductive Sciences, University of California, San Francisco, United States.
Balázs
B
Legeza
Department of Obstetrics and Gynecology, Virginia Commonwealth University, Richmond, VA 23298, United States; Department of Human and Molecular Genetics, Virginia Commonwealth University, Richmond, VA 23298, United States.
Bhavi
B
Modi
Department of Obstetrics and Gynecology, Virginia Commonwealth University, Richmond, VA 23298, United States; Department of Human and Molecular Genetics, Virginia Commonwealth University, Richmond, VA 23298, United States.
Maria Eugenia
ME
Teves
Department of Pathology, Pennsylvania State College of Medicine, Hershey, PA 17033, United States; Department of Obstetrics and Gynecology, Pennsylvania State College of Medicine, Hershey, PA 17033, United States.
Janette M
JM
McAllister
Department of Obstetrics and Gynecology, Virginia Commonwealth University, Richmond, VA 23298, United States; Department of Human and Molecular Genetics, Virginia Commonwealth University, Richmond, VA 23298, United States.
Jerome F
JF
Strauss
Department of Pediatrics and the Center for Reproductive Sciences, University of California, San Francisco, United States. Electronic address: wlmlab@ucsf.edu.
Walter L
WL
Miller
D017398:Alternative Splicing; D002460:Cell Line; D002786:Cholesterol Side-Chain Cleavage Enzyme / Q000235:genetics; D053418:Death Domain Receptor Signaling Adaptor Proteins / Q000235:genetics*; D005260:Female; D014644:Genetic Variation; D055106:Genome-Wide Association Study; D020662:Guanine Nucleotide Exchange Factors / Q000235:genetics*; D006801:Humans; D011085:Polycystic Ovary Syndrome / Q000235:genetics*; D017422:Sequence Analysis, DNA; D013254:Steroid 17-alpha-Hydroxylase / Q000235:genetics; D013799:Theca Cells / Q000737:chemistry*; D015854:Up-Regulation
C571984:DENND1A protein, human; D053418:Death Domain Receptor Signaling Adaptor Proteins; D020662:Guanine Nucleotide Exchange Factors; C501892:CYP17A1 protein, human; D013254:Steroid 17-alpha-Hydroxylase; D002786:Cholesterol Side-Chain Cleavage Enzyme
Adrenal; Alternate splicing; Androgens; NCI-H295A cells; Ovary; RNA splicing