title: Actin polymerization in differentiated vascular smooth muscle cells requires vasodilator-stimulated phosphoprotein.
abstract: Our group has previously shown that vasoconstrictors increase net actin polymerization in differentiated vascular smooth muscle cells (dVSMC) and that increased actin polymerization is linked to contractility of vascular tissue (Kim et al., Am J Physiol Cell Physiol 295: C768-778, 2008). However, the underlying mechanisms are largely unknown. Here, we evaluated the possible functions of the Ena/vasodilator-stimulated phosphoprotein (VASP) family of actin filament elongation factors in dVSMC. Inhibition of actin filament elongation by cytochalasin D decreases contractility without changing myosin light-chain phosphorylation levels, suggesting that actin filament elongation is necessary for dVSM contraction. VASP is the only Ena/VASP protein highly expressed in aorta tissues, and VASP knockdown decreased smooth muscle contractility. VASP partially colocalizes with alpha-actinin and vinculin in dVSMC. Profilin, known to associate with G actin and VASP, also colocalizes with alpha-actinin and vinculin, potentially identifying the dense bodies and the adhesion plaques as hot spots of actin polymerization. The EVH1 domain of Ena/VASP is known to target these proteins to their sites of action. Introduction of an expressed EVH1 domain as a dominant negative inhibits stimulus-induced increases in actin polymerization. VASP phosphorylation, known to inhibit actin polymerization, is decreased during phenylephrine stimulation in dVSMC. We also directly visualized, for the first time, rhodamine-labeled actin incorporation in dVSMC and identified hot spots of actin polymerization in the cell cortex that colocalize with VASP. These results indicate a role for VASP in actin filament assembly, specifically at the cell cortex, that modulates contractility in dVSMC.
authors.affiliation: Dept. of Health Sciences, Boston Univ., 635 Commonwealth Ave, Boston MA 02215, USA.
authors.forename: Hak Rim
authors.initials: HR
authors.lastname: Kim
authors.forename: Philip
authors.initials: P
authors.lastname: Graceffa
authors.forename: François
authors.initials: F
authors.lastname: Ferron
authors.forename: Cynthia
authors.initials: C
authors.lastname: Gallant
authors.forename: Malgorzata
authors.initials: M
authors.lastname: Boczkowska
authors.forename: Roberto
authors.initials: R
authors.lastname: Dominguez
authors.forename: Kathleen G
authors.initials: KG
authors.lastname: Morgan
mesh_terms: D000185:Actinin / Q000378:metabolism; D000199:Actins / Q000378:metabolism*; D000316:Adrenergic alpha-Agonists / Q000494:pharmacology; D000595:Amino Acid Sequence; D000818:Animals; D001011:Aorta / Q000378:metabolism; D015815:Cell Adhesion Molecules / Q000235:genetics / Q000378:metabolism*; D002454:Cell Differentiation / Q000187:drug effects; D015638:Cytochalasin D / Q000494:pharmacology; D005289:Ferrets; D006801:Humans; D008840:Microfilament Proteins / Q000235:genetics / Q000378:metabolism*; D008856:Microscopy, Fluorescence; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009131:Muscle, Smooth, Vascular / Q000187:drug effects / Q000378:metabolism*; D009154:Mutation; D032389:Myocytes, Smooth Muscle / Q000187:drug effects / Q000378:metabolism*; D016376:Oligonucleotides, Antisense / Q000378:metabolism; D009924:Organ Culture Techniques; D010656:Phenylephrine / Q000494:pharmacology; D010750:Phosphoproteins / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D051304:Profilins / Q000378:metabolism; D055503:Protein Multimerization; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D015398:Signal Transduction; D013997:Time Factors; D014661:Vasoconstriction / Q000187:drug effects; D014662:Vasoconstrictor Agents / Q000494:pharmacology; D016596:Vinculin / Q000378:metabolism; D000098662:Vasodilator-Stimulated Phosphoprotein
chemical_list: D000199:Actins; D000316:Adrenergic alpha-Agonists; D015815:Cell Adhesion Molecules; C484449:EVL protein, human; D008840:Microfilament Proteins; D016376:Oligonucleotides, Antisense; D010750:Phosphoproteins; D051304:Profilins; D011993:Recombinant Fusion Proteins; D014662:Vasoconstrictor Agents; D000098662:Vasodilator-Stimulated Phosphoprotein; D000185:Actinin; D016596:Vinculin; D010656:Phenylephrine; D015638:Cytochalasin D