Neoinitimal hyperplasia is a common complicating feature of atherosclerosis, restenosis after angioplasty, and transplant vasculopathy. At the cell level, over-growth of the arterial wall is triggered by inflammatory signals that promote progressive phenotypic changes resulting in enhanced cell proliferation and migration. Evidence suggests that this dedifferentiation process known as phenotypic modulation may occur in both medial smooth muscle cells and adventitial fibroblasts where the biochemical hallmark of cell reprogramming is alteration in expression of vascular smooth muscle (VSM) alpha-actin, a cytoskeletal protein important for regulation of vascular contraction and cell movement. Hence, elucidation of the mechanisms governing expression of the VSM alpha-actin gene in cells of myogenic and fibroblastic lineage may reveal molecular targets of clinical utility in the management of atherosclerotic plaque stability and/or restenosis after surgical intervention. The primary goal of this research is to define the gene regulatory function(s) of a novel group of single-stranded DNA and mRNA-binding factors known as Puralpha, Purbeta, and MSYI. These proteins have been implicated in coordinately suppressing VSM alpha-actin gene expression in myofibroblasts and smooth muscle cells by both transcriptional and post-transcriptional mechanisms. In this proposal, particular emphasis will be placed on the participation of Purbeta in these processes since this protein appears to be the key component required for repression of the VSM alpha-actin promoter in cultured cells and because its pattern of expression in vascular tissue specimens is consistent with a role in phenotypic modulation. In vitro and in vivo approaches will be used to test predictions arising from conceptual models of gene regulation by 1) mapping structural domains and chemical modifications required for interaction of Purbeta with relevant nucleic acid and protein-binding partners 2) analyzing the assembly and disassembly of Pur-containing regulatory complexes on the genomic VSM alpha-actin promoter, 3) evaluating the ability of Pur and Y-box proteins to inhibit translation of VSM alpha-actin mRNA, and 4) characterizing the time-course of expression of Puralpha, Purbeta, and MSY1 in mouse models of vascular disease and injury. This research will contribute to an improved understanding of a unique class of single-stranded nucleic acid-binding factors that may account for the remarkable plasticity of VSM alpha-actin expression and phenotypic modulation during vascular remodeling.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL054281-22
Application #
6780918
Study Section
Experimental Cardiovascular Sciences Study Section (ECS)
Program Officer
Ershow, Abby
Project Start
1995-08-01
Project End
2007-07-31
Budget Start
2004-08-01
Budget End
2005-07-31
Support Year
22
Fiscal Year
2004
Total Cost
$264,652
Indirect Cost
Name
University of Vermont & St Agric College
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
066811191
City
Burlington
State
VT
Country
United States
Zip Code
05405
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David, Jason J; Subramanian, Sukanya V; Zhang, Aiwen et al. (2012) Y-box binding protein-1 implicated in translational control of fetal myocardial gene expression after cardiac transplant. Exp Biol Med (Maywood) 237:593-607
Rumora, Amy E; Steere, Ashley N; Ramsey, Jon E et al. (2010) Isolation and characterization of the core single-stranded DNA-binding domain of purine-rich element binding protein B (Purýý). Biochem Biophys Res Commun 400:340-5
Zhao, Sheng; Kelm Jr, Robert J; Fernald, Russell D (2010) Regulation of gonadotropin-releasing hormone-1 gene transcription by members of the purine-rich element-binding protein family. Am J Physiol Endocrinol Metab 298:E524-33
Liu, Xiaoying; Kelm Jr, Robert J; Strauch, Arthur R (2009) Transforming growth factor beta1-mediated activation of the smooth muscle alpha-actin gene in human pulmonary myofibroblasts is inhibited by tumor necrosis factor-alpha via mitogen-activated protein kinase kinase 1-dependent induction of the Egr-1 transcr Mol Biol Cell 20:2174-85
Ramsey, Jon E; Kelm Jr, Robert J (2009) Mechanism of strand-specific smooth muscle alpha-actin enhancer interaction by purine-rich element binding protein B (Purbeta). Biochemistry 48:6348-60
Zhang, Aiwen; David, Jason J; Subramanian, Sukanya V et al. (2008) Serum response factor neutralizes Pur alpha- and Pur beta-mediated repression of the fetal vascular smooth muscle alpha-actin gene in stressed adult cardiomyocytes. Am J Physiol Cell Physiol 294:C702-14
Ramsey, Jon E; Daugherty, Margaret A; Kelm Jr, Robert J (2007) Hydrodynamic studies on the quaternary structure of recombinant mouse Purbeta. J Biol Chem 282:1552-60
Ji, Juan; Tsika, Gretchen L; Rindt, Hansjorg et al. (2007) Puralpha and Purbeta collaborate with Sp3 to negatively regulate beta-myosin heavy chain gene expression during skeletal muscle inactivity. Mol Cell Biol 27:1531-43
Knapp, Anna M; Ramsey, Jon E; Wang, Shu-Xia et al. (2007) Structure-function analysis of mouse Pur beta II. Conformation altering mutations disrupt single-stranded DNA and protein interactions crucial to smooth muscle alpha-actin gene repression. J Biol Chem 282:35899-909

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