A novel β-adrenergic response element regulates both basal and agonist-induced expression of cyclin-dependent kinase 1 gene in cardiac fibroblasts.
Cardiac fibrosis, a known risk factor for heart disease, is typically caused by uncontrolled proliferation of fibroblasts and excessive deposition of extracellular matrix proteins in the myocardium. Cyclin-dependent kinase 1 (CDK1) is involved in the control of G2/M transit phase of the cell cycle. Here, we showed that isoproterenol (ISO)-induced cardiac fibrosis is associated with increased levels of CDK1 exclusively in fibroblasts in the adult mouse heart. Treatment of primary embryonic ventricular cell cultures with ISO (a nonselective β-adrenergic receptor agonist) increased CDK1 protein expression in fibroblasts and promoted their cell cycle activity. Quantitative PCR analysis confirmed that ISO increases CDK1 transcription in a transient manner. Further, the ISO-responsive element was mapped to the proximal -100-bp sequence of the CDK1 promoter region using various 5'-flanking sequence deletion constructs. Sequence analysis of the -100-bp CDK1 minimal promoter region revealed two putative nuclear factor-Y (NF-Y) binding elements. Overexpression of the NF-YA subunit in primary ventricular cultures significantly increased the basal activation of the -100-bp CDK1 promoter construct but not the ISO-induced transcription of the minimal promoter construct. In contrast, dominant negative NF-YA expression decreased the basal activity of the minimal promoter construct and ISO treatment fully rescued the dominant negative effects. Furthermore, site-directed mutagenesis of the distal NF-Y binding site in the -100-bp CDK1 promoter region completely abolished both basal and ISO-induced promoter activation of the CDK1 gene. Collectively, our results raise an exciting possibility that targeting CDK1 or NF-Y in the diseased heart may inhibit fibrosis and subsequently confer cardioprotection.
Department of Pharmacology, Dalhousie University, Halifax, Nova Scotia, Canada.
Gerard J
GJ
Gaspard
Jessica
J
MacLean
Danielle
D
Rioux
Kishore B S
KB
Pasumarthi
D000318:Adrenergic beta-Agonists / Q000494:pharmacology; D000818:Animals; D023081:CCAAT-Binding Factor / Q000235:genetics / Q000378:metabolism*; D016203:CDC2 Protein Kinase / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D049109:Cell Proliferation; D002478:Cells, Cultured; D004268:DNA-Binding Proteins / Q000235:genetics; D005347:Fibroblasts / Q000378:metabolism; D005355:Fibrosis; D007545:Isoproterenol; D008297:Male; D051379:Mice; D009206:Myocardium / Q000201:enzymology* / Q000473:pathology*; D011401:Promoter Regions, Genetic / Q000235:genetics; D012333:RNA, Messenger / Q000096:biosynthesis; D020218:Response Elements
D000318:Adrenergic beta-Agonists; D023081:CCAAT-Binding Factor; D004268:DNA-Binding Proteins; C492060:Nfya protein, mouse; D012333:RNA, Messenger; D016203:CDC2 Protein Kinase; D007545:Isoproterenol
CDK1; NF-Y; cardiac fibroblast proliferation; isoproterenol; β-adrenergic receptor agonist