Gap junctions in Vascular Smooth Muscle: growth control

Summary

Principal Investigator: Janis M Burt
Abstract: Project Description The long-term goals of this research are: 1) to understand the molecular mechanisms that govern the permeability and growth suppressive properties of vascular cell gap junctions and 2) to develop from that knowledge the rationale for gene therapies that target connexin expression in endothelial cells with the goal of limiting susceptibility to and facilitating recovery from ischemic injury. Three gap junction proteins are commonly expressed in vascular cells, connexin (Cx) 37, Cx40 and Cx43;in the endothelium Cx37 and Cx40 normally predominate, but during vasculogenesis and with stress, injury and disease, Cx37 is down-regulated and Cx43 up-regulated. The consequences of this change in expression on the vessel's ability to form new vessels and to maintain vessel functions during vascular remodeling remain uncertain. In our previous studies, we demonstrated that these connexins form gap junction channels with vastly different permselective and growth suppressive properties that are regulated by growth factor activated signaling cascades in a connexin- specific manner. In the current proposal we hypothesize that connexin-specific, phosphorylation-dependent regulation of junctional permselectivity provides vascular cells a strategy for maintaining coordinated contraction/relaxation functions of vessels while simultaneously supporting the proliferative response of cells therein. We address this hypothesis in Aim 1 by examining the mechanistic basis for how phosphorylation events in the carboxyl terminal domain (CT) lead to altered permselective properties of the associated pore domain. In aim 2 we extend the observations of Aim 1 and determine whether the growth suppressive properties of these connexins rely on their permselective properties and/or their direct interactions with proteins involved in cell cycle control and progression. These growth studies make use of Cx-deficient cell lines, endothelial cells isolated from wild type or Cx37 deficient mice, and an in vivo hindlimb ischemic injury model, asking whether Cx37 works in conjunction with or in opposition to Cx43 to regulate the angiogenic response induced by injury while preserving junctional permselective properties. A combination of electrophysiology and fluorescence microscopy will be used to quantify the permselective properties of junctions and molecular approaches will be used to identify essential regions/sites of interaction between connexin domains and between connexins and elements of the cell cycle machinery. Isolated endothelial cells and an in vivo ischemia model will be used to determine the benefit of connexin expression or silencing to the extent and speed of vascular remodeling following injury. Our studies can be expected to lend new insights on the mechanistic basis for phosphorylation-dependent regulation of the permeability and growth suppressive functions of the vascular connexins and to the possible use of gene therapy to manipulate connexin expression in the endothelium to maximize/minimize angiogenesis, as appropriate, in settings of vascular injury and disease.
Funding Period: 1997-07-15 - 2014-11-30
more information: NIH RePORT

Top Publications

  1. pmc Selectivity of connexin 43 channels is regulated through protein kinase C-dependent phosphorylation
    Jose F Ek-Vitorin
    Department of Physiology, University of Arizona, Tucson 85724, USA
    Circ Res 98:1498-505. 2006
  2. pmc An electrically coupled tissue-engineered cardiomyocyte scaffold improves cardiac function in rats with chronic heart failure
    Jordan J Lancaster
    Cardiology and Medicine, Southern Arizona VA Health Care System Sarver Heart Center Department of Physiology Electronic address
    J Heart Lung Transplant 33:438-45. 2014
  3. pmc Carboxy terminus and pore-forming domain properties specific to Cx37 are necessary for Cx37-mediated suppression of insulinoma cell proliferation
    Tasha K Nelson
    Department of Physiology, University of Arizona, Tucson, Arizona and
    Am J Physiol Cell Physiol 305:C1246-56. 2013
  4. pmc Compromised regulation of tissue perfusion and arteriogenesis limit, in an AT1R-independent fashion, recovery of ischemic tissue in Cx40(-/-) mice
    Jennifer S Fang
    Department of Physiology, University of Arizona, Tucson, AZ 85718, USA
    Am J Physiol Heart Circ Physiol 304:H816-27. 2013
  5. pmc Extracellular loop cysteine mutant of cx37 fails to suppress proliferation of rat insulinoma cells
    Miranda E Good
    Department of Physiology, University of Arizona, PO Box 245051, Tucson, AZ 85724, USA
    J Membr Biol 245:369-80. 2012
  6. pmc Inducible coexpression of connexin37 or connexin40 with connexin43 selectively affects intercellular molecular transfer
    Joanna Gemel
    Department of Pediatrics, University of Chicago, Chicago, IL 60637, USA
    J Membr Biol 245:231-41. 2012
  7. pmc Intercellular calcium waves in primary cultured rat mesenteric smooth muscle cells are mediated by connexin43
    Nadia Halidi
    Laboratory of Cell Biophysics, Ecole Polytechnique Federale de Lausanne, Switzerland
    Cell Commun Adhes 19:25-37. 2012
  8. pmc Structural basis for the selective permeability of channels made of communicating junction proteins
    Jose F Ek-Vitorin
    Department of Physiology, University of Arizona, Tucson, AZ, USA
    Biochim Biophys Acta 1828:51-68. 2013
  9. pmc Cx40 is required for, and cx37 limits, postischemic hindlimb perfusion, survival and recovery
    Jennifer S Fang
    Department of Physiology, University of Arizona Health Sciences Center, Tucson, Ariz, USA
    J Vasc Res 49:2-12. 2012
  10. pmc Cx37 deletion enhances vascular growth and facilitates ischemic limb recovery
    Jennifer S Fang
    Department of Physiology, University of Arizona, Tucson, Arizona, USA
    Am J Physiol Heart Circ Physiol 301:H1872-81. 2011

Research Grants

  1. RAGE and Mechanisms of Vascular Dysfunction
    Shi Fang Yan; Fiscal Year: 2013

Detail Information

Publications16

  1. pmc Selectivity of connexin 43 channels is regulated through protein kinase C-dependent phosphorylation
    Jose F Ek-Vitorin
    Department of Physiology, University of Arizona, Tucson 85724, USA
    Circ Res 98:1498-505. 2006
    ..Our data suggest that the selectivity of gap junction channels at intercalated disks is increased early in ischemia...
  2. pmc An electrically coupled tissue-engineered cardiomyocyte scaffold improves cardiac function in rats with chronic heart failure
    Jordan J Lancaster
    Cardiology and Medicine, Southern Arizona VA Health Care System Sarver Heart Center Department of Physiology Electronic address
    J Heart Lung Transplant 33:438-45. 2014
    ..This study examines an angiogenic and biodegradable cardiac construct seeded with neonatal cardiomyocytes for the treatment of chronic heart failure (CHF)...
  3. pmc Carboxy terminus and pore-forming domain properties specific to Cx37 are necessary for Cx37-mediated suppression of insulinoma cell proliferation
    Tasha K Nelson
    Department of Physiology, University of Arizona, Tucson, Arizona and
    Am J Physiol Cell Physiol 305:C1246-56. 2013
    ....
  4. pmc Compromised regulation of tissue perfusion and arteriogenesis limit, in an AT1R-independent fashion, recovery of ischemic tissue in Cx40(-/-) mice
    Jennifer S Fang
    Department of Physiology, University of Arizona, Tucson, AZ 85718, USA
    Am J Physiol Heart Circ Physiol 304:H816-27. 2013
    ..Our data suggest that Cx40(-/-) mice are at significantly greater risk for poor recovery from ischemic insult due to compromised regulation of tissue perfusion, vascular remodeling, and prolonged inflammatory response...
  5. pmc Extracellular loop cysteine mutant of cx37 fails to suppress proliferation of rat insulinoma cells
    Miranda E Good
    Department of Physiology, University of Arizona, PO Box 245051, Tucson, AZ 85724, USA
    J Membr Biol 245:369-80. 2012
    ....
  6. pmc Inducible coexpression of connexin37 or connexin40 with connexin43 selectively affects intercellular molecular transfer
    Joanna Gemel
    Department of Pediatrics, University of Chicago, Chicago, IL 60637, USA
    J Membr Biol 245:231-41. 2012
    ..These data show that induced expression of either Cx37 or Cx40 in Cx43-expressing cells can selectively alter the intercellular exchange of some molecules without affecting the transfer of others...
  7. pmc Intercellular calcium waves in primary cultured rat mesenteric smooth muscle cells are mediated by connexin43
    Nadia Halidi
    Laboratory of Cell Biophysics, Ecole Polytechnique Federale de Lausanne, Switzerland
    Cell Commun Adhes 19:25-37. 2012
    ....
  8. pmc Structural basis for the selective permeability of channels made of communicating junction proteins
    Jose F Ek-Vitorin
    Department of Physiology, University of Arizona, Tucson, AZ, USA
    Biochim Biophys Acta 1828:51-68. 2013
    ..This article is part of a Special Issue entitled: The Communicating junctions, roles and dysfunctions...
  9. pmc Cx40 is required for, and cx37 limits, postischemic hindlimb perfusion, survival and recovery
    Jennifer S Fang
    Department of Physiology, University of Arizona Health Sciences Center, Tucson, Ariz, USA
    J Vasc Res 49:2-12. 2012
    ..Vascular endothelium predominantly expresses two connexin (Cx) isoforms: Cx37 and Cx40. The relevance of these Cxs to postischemic limb recovery remains unclear...
  10. pmc Cx37 deletion enhances vascular growth and facilitates ischemic limb recovery
    Jennifer S Fang
    Department of Physiology, University of Arizona, Tucson, Arizona, USA
    Am J Physiol Heart Circ Physiol 301:H1872-81. 2011
    ..These results are consistent with Cx37 exerting a growth-suppressive effect in the vasculature that limits embryonic vasculogenesis as well as arteriogenic and angiogenic responses to ischemic injury in the adult animal...
  11. pmc A functional channel is necessary for growth suppression by Cx37
    Miranda E Good
    Department of Physiology, University of Arizona, PO Box 245051, Tucson, AZ 85724, USA
    J Cell Sci 124:2448-56. 2011
    ....
  12. pmc Regulation of gap junctional charge selectivity in cells coexpressing connexin 40 and connexin 43
    Nathanael S Heyman
    Department of Physiology, University of Arizona, Tucson, AZ 85724, USA
    Am J Physiol Heart Circ Physiol 297:H450-9. 2009
    ..Such regulation could represent a mechanism by which coexpressing cells such as vascular endothelium and atrial cells regulate acutely the selective intercellular communication mediated by their gap junctions...
  13. pmc Connexin 37 profoundly slows cell cycle progression in rat insulinoma cells
    Janis M Burt
    Dept of Physiology, P O Box 245051, Univ of Arizona, Tucson, AZ 85724, USA
    Am J Physiol Cell Physiol 295:C1103-12. 2008
    ..Our results indicate that Cx37 expression suppresses cell proliferation by significantly increasing cell cycle time by extending all phases of the cell cycle and accumulating cells at the G(1)/S checkpoint...
  14. pmc Hindered diffusion through an aqueous pore describes invariant dye selectivity of Cx43 junctions
    Nathanael S Heyman
    Department of Physiology, University of Arizona, Tucson, Arizona 85724, USA
    Biophys J 94:840-54. 2008
    ..The permselectivity and dye selectivity data suggest the variable presence in Cx43-comprised junctions of conductive channels that are either dye-impermeable or dye-permeable...
  15. pmc Analysis of Connexin43 phosphorylated at S325, S328 and S330 in normoxic and ischemic heart
    Paul D Lampe
    Molecular Diagnostics Program, Fred Hutchinson Cancer Research Center, Department of Pathobiology, University of Washington, 1100 Fairview Avenue N, M5C800, PO Box 19024, Seattle, WA 98109, USA
    J Cell Sci 119:3435-42. 2006
    ..These data suggest that phosphorylation of Cx43 at serine residues 325, 328 and/or 330 influences channel permselectivity and regulates the efficiency of gap junction assembly...
  16. pmc Mix and match: investigating heteromeric and heterotypic gap junction channels in model systems and native tissues
    Michael Koval
    Division of Pulmonary, Allergy and Critical Care Medicine, Department of Medicine, Emory University, Atlanta, GA, United States Department of Cell Biology, Emory University, Atlanta, GA, United States Electronic address
    FEBS Lett 588:1193-204. 2014
    ..There remains a need to use and develop different experimental approaches in order to understand the prevalence and roles for mixed gap junction channels in human physiology. ..

Research Grants30

  1. RAGE and Mechanisms of Vascular Dysfunction
    Shi Fang Yan; Fiscal Year: 2013
    ..Using novel and state-of-the-art techniques, floxed mice and molecular approaches to gene regulation, we are well-positioned to lead the study of RAGE in the next cycle of this Program. ..