title: Nicotine induced CpG methylation of Pax6 binding motif in StAR promoter reduces the gene expression and cortisol production.
abstract: Steroidogenic acute regulatory protein (StAR) mediates the rate-limiting step in the synthesis of steroid hormones, essential to fetal development. We have reported that the StAR expression in fetal adrenal is inhibited in a rat model of nicotine-induced intrauterine growth retardation (IUGR). Here using primary human fetal adrenal cortex (pHFAC) cells and a human fetal adrenal cell line NCI-H295A, we show that nicotine inhibits StAR expression and cortisol production in a dose- and time-dependent manner, and prolongs the inhibitory effect on cells proliferating over 5 passages after termination of nicotine treatment. Methylation detection within the StAR promoter region uncovers a single site CpG methylation at nt -377 that is sensitive to nicotine treatment. Nicotine-induced alterations in frequency of this point methylation correlates well with the levels of StAR expression, suggesting an important role of the single site in regulating StAR expression. Further studies using bioinformatics analysis and siRNA approach reveal that the single CpG site is part of the Pax6 binding motif (CGCCTGA) in the StAR promoter. The luciferase activity assays validate that Pax6 increases StAR gene expression by binding to the glucagon G3-like motif (CGCCTGA) and methylation of this site blocks Pax6 binding and thus suppresses StAR expression. These data identify a nicotine-sensitive CpG site at the Pax6 binding motif in the StAR promoter that may play a central role in regulating StAR expression. The results suggest an epigenetic mechanism that may explain how nicotine contributes to onset of adult diseases or disorders such as metabolic syndrome via fetal programming.
authors.affiliation: Department of Pharmacology, Basic Medical School of Wuhan University, Wuhan 430071, China.
authors.forename: Tingting
authors.initials: T
authors.lastname: Wang
authors.forename: Man
authors.initials: M
authors.lastname: Chen
authors.forename: Lian
authors.initials: L
authors.lastname: Liu
authors.forename: Huaiyan
authors.initials: H
authors.lastname: Cheng
authors.forename: You-E
authors.initials: YE
authors.lastname: Yan
authors.forename: Ying-Hong
authors.initials: YH
authors.lastname: Feng
authors.forename: Hui
authors.initials: H
authors.lastname: Wang
mesh_terms: D000302:Adrenal Cortex / Q000187:drug effects / Q000196:embryology / Q000378:metabolism; D020816:Amino Acid Motifs; D002460:Cell Line; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D018899:CpG Islands / Q000235:genetics*; D019175:DNA Methylation / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D005136:Eye Proteins / Q000235:genetics*; D018507:Gene Expression Regulation, Developmental / Q000187:drug effects; D005934:Glucagon / Q000235:genetics; D018398:Homeodomain Proteins / Q000235:genetics*; D006801:Humans; D006854:Hydrocortisone / Q000378:metabolism; D009538:Nicotine / Q000008:administration & dosage / Q000633:toxicity*; D018722:Nicotinic Agonists / Q000008:administration & dosage / Q000633:toxicity*; D000071841:PAX6 Transcription Factor; D051761:Paired Box Transcription Factors / Q000235:genetics*; D010750:Phosphoproteins / Q000235:genetics*; D011401:Promoter Regions, Genetic; D012097:Repressor Proteins / Q000235:genetics*; D013997:Time Factors; D000098682:Steroidogenic Acute Regulatory Protein
chemical_list: D005136:Eye Proteins; D018398:Homeodomain Proteins; D018722:Nicotinic Agonists; D000071841:PAX6 Transcription Factor; C000605730:PAX6 protein, human; D051761:Paired Box Transcription Factors; C000605733:Pax6 protein, rat; D010750:Phosphoproteins; D012097:Repressor Proteins; D000098682:Steroidogenic Acute Regulatory Protein; D009538:Nicotine; D005934:Glucagon; D006854:Hydrocortisone