title: ERK1/2-dependent contractile protein expression in vascular smooth muscle cells.
abstract: In vivo, vascular smooth muscle (VSM) cells change their contractile phenotype toward a more proliferative phenotype during the pathogenesis of vascular diseases. Because these dedifferentiated VSM cells may gradually regain contractile functions, we aimed to identify signaling pathways that result in an increased expression of contractile proteins in VSM cells. In vitro, serum and thrombin induced a reversible upregulation of smooth muscle myosin heavy-chain (SM-MHC) in cultured neonatal rat VSM cells. Cotransfection of a SM-MHC-promoter chloramphenicol acetyltransferase-construct with dominant-negative N17Ras or N17Raf or treatment with the mitogen-activated/ERK-activating kinase (MEK) inhibitor PD 98059 concentration dependently decreased the serum- or thrombin-induced SM-MHC promoter activity. Consistently, the serum- or thrombin-induced phosphorylation of MEK and extracellular signal-regulated kinase 1/2 (ERK1/2) coincided with a MEK-dependent nuclear accumulation of phosphorylated ERK1/2 and subsequent nuclear phosphorylation of the transcription factors c-myc and Elk-1. A 5'-deletion analysis of cis-elements within the SM-MHC promoter demonstrated that a conserved region (nucleotide -1346 to -1102) was required for both cell type-specific expression and serum- or thrombin-induced upregulation of the SM-MHC promoter in VSM cells. Within this region, 2 CArG-boxes, a GC-rich element, and a CTF/NF-1 site are critical positively acting cis-elements for the serum- or thrombin-induced upregulation of SM-MHC. We conclude that the serum- or thrombin-induced differentiation requires an intact Ras/Raf/MEK/ERK signaling cascade, nuclear translocation of activated ERK1/2, phosphorylation of transcription factors, and several cis-elements within the SM-MHC promoter.
authors.affiliation: Institut für Klinische Pharmakologie und Toxikologie, Freie Universität Berlin, Berlin, Germany.
authors.forename: Dag
authors.initials: D
authors.lastname: Schauwienold
authors.forename: Claudia
authors.initials: C
authors.lastname: Plum
authors.forename: Thomas
authors.initials: T
authors.lastname: Helbing
authors.forename: Philipp
authors.initials: P
authors.lastname: Voigt
authors.forename: Thomas
authors.initials: T
authors.lastname: Bobbert
authors.forename: Dorett
authors.initials: D
authors.lastname: Hoffmann
authors.forename: Martin
authors.initials: M
authors.lastname: Paul
authors.forename: Hans Peter
authors.initials: HP
authors.lastname: Reusch
mesh_terms: D000818:Animals; D002467:Cell Nucleus / Q000378:metabolism; D002478:Cells, Cultured; D003470:Culture Media / Q000494:pharmacology; D004268:DNA-Binding Proteins; D020935:MAP Kinase Signaling System; D019950:Mitogen-Activated Protein Kinase 1 / Q000502:physiology*; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases / Q000502:physiology*; D009131:Muscle, Smooth, Vascular / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D018995:Myosin Heavy Chains / Q000096:biosynthesis / Q000235:genetics*; D010766:Phosphorylation; D011401:Promoter Regions, Genetic; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D016271:Proto-Oncogene Proteins c-myc / Q000378:metabolism; D012333:RNA, Messenger / Q000096:biosynthesis; D051381:Rats; D020218:Response Elements; D013917:Thrombin / Q000494:pharmacology; D014157:Transcription Factors; D015533:Transcriptional Activation; D050807:ets-Domain Protein Elk-1
chemical_list: D003470:Culture Media; D004268:DNA-Binding Proteins; C494114:Elk1 protein, rat; D011518:Proto-Oncogene Proteins; D016271:Proto-Oncogene Proteins c-myc; D012333:RNA, Messenger; D014157:Transcription Factors; D050807:ets-Domain Protein Elk-1; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases; D013917:Thrombin; D018995:Myosin Heavy Chains