title: Transforming growth factor-beta1 (TGF-beta1) utilizes distinct pathways for the transcriptional activation of microRNA 143/145 in human coronary artery smooth muscle cells.
abstract: MicroRNA 143/145 (miR143/145) is restricted to adult smooth muscle cell (SMC) lineages and mediates, in part, the expression of several SMC contractile genes. Although the function of miR143/145 has begun to be elucidated, its transcriptional regulation in response to various signaling inputs is poorly understood. In an effort to define a miR signature for SMC differentiation, we screened human coronary artery SMCs for miRs modulated by TGF-β1, a known stimulus for SMC differentiation. Array analysis revealed a number of TGF-β1-induced miRs, including miR143/145. Validation studies showed that TGF-β1 stimulated miR143/145 expression in a dose- and time-dependent manner. We utilized several chemical inhibitors and found that SB203580, a specific inhibitor of p38MAPK, significantly decreased TGF-β1-induced miR143/145 expression. siRNA studies demonstrated that the effect of TGF-β1 on miR143/145 was dependent upon the myocardin and serum response factor transcriptional switch as well as SMAD4. TGF-β1 stimulated a 580-bp human miR143/145 enhancer, and mutagenesis studies revealed a critical role for both a known CArG box and an adjacent SMAD-binding element for full TGF-β1-dependent activation of the enhancer. Chromatin immunoprecipitation assays documented TGF-β1-mediated enrichment of SMAD3 and SMAD4 binding over the enhancer region containing the SMAD-binding element. Pre-miR145 strongly promoted SMC differentiation, whereas an anti-miR145 partially blocked TGF-β1-induced SMC differentiation. These results demonstrate a dual pathway for TGF-β1-induced transcription of miR143/145, thus revealing a novel mechanism underlying TGF-β1-induced human vascular SMC differentiation.
authors.affiliation: Aab Cardiovascular Research Institute, Department of Medicine, University of Rochester School of Medicine and Dentistry, Rochester, New York 14642, USA. xiaochun_long@urmc.rochester.edu
authors.forename: Xiaochun
authors.initials: X
authors.lastname: Long
authors.forename: Joseph M
authors.initials: JM
authors.lastname: Miano
mesh_terms: D000818:Animals; D002454:Cell Differentiation / Q000187:drug effects / Q000502:physiology; D002478:Cells, Cultured; D003331:Coronary Vessels / Q000166:cytology / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005786:Gene Expression Regulation / Q000187:drug effects / Q000502:physiology; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology; D035683:MicroRNAs / Q000096:biosynthesis* / Q000235:genetics; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000378:metabolism*; D032389:Myocytes, Smooth Muscle / Q000166:cytology / Q000378:metabolism*; D011725:Pyridines / Q000494:pharmacology; D051381:Rats; D020218:Response Elements / Q000502:physiology; D051900:Smad3 Protein / Q000235:genetics / Q000378:metabolism; D051901:Smad4 Protein / Q000235:genetics / Q000378:metabolism; D014158:Transcription, Genetic / Q000502:physiology*; D053773:Transforming Growth Factor beta1 / Q000378:metabolism / Q000494:pharmacology*; D048051:p38 Mitogen-Activated Protein Kinases / Q000235:genetics / Q000378:metabolism
chemical_list: D004791:Enzyme Inhibitors; D007093:Imidazoles; C542359:MIRN143 microRNA, human; C541010:MIRN145 microRNA, human; C542511:MIRN145 microRNA, rat; D035683:MicroRNAs; D011725:Pyridines; C497790:SMAD3 protein, human; C497796:SMAD4 protein, human; D051900:Smad3 Protein; C497787:Smad3 protein, rat; D051901:Smad4 Protein; C490811:Smad4 protein, rat; D053773:Transforming Growth Factor beta1; D048051:p38 Mitogen-Activated Protein Kinases; C093642:SB 203580