title: Stimulus-specific activation and actin dependency of distinct, spatially separated ERK1/2 fractions in A7r5 smooth muscle cells.
abstract: A proliferative response of smooth muscle cells to activation of extracellular signal regulated kinases 1 and 2 (ERK1/2) has been linked to cardiovascular disease. In fully differentiated smooth muscle, however, ERK1/2 activation can also regulate contraction. Here, we use A7r5 smooth muscle cells, stimulated with 12-deoxyphorbol 13-isobutylate 20-acetate (DPBA) to induce cytoskeletal remodeling or fetal calf serum (FCS) to induce proliferation, to identify factors that determine the outcomes of ERK1/2 activation in smooth muscle. Knock down experiments, immunoprecipitation and proximity ligation assays show that the ERK1/2 scaffold caveolin-1 mediates ERK1/2 activation in response to DPBA, but not FCS, and that ERK1/2 is released from caveolin-1 upon DPBA, but not FCS, stimulation. Conversely, ERK1/2 associated with the actin cytoskeleton is significantly reduced after FCS, but not DPBA stimulation, as determined by Triton X fractionation. Furthermore, cytochalasin treatment inhibits DPBA, but not FCS-induced ERK1/2 phosphorylation, indicating that the actin cytoskeleton is not only a target but also is required for ERK1/2 activation. Our results show that (1) at least two ERK1/2 fractions are regulated separately by specific stimuli, and that (2) the association of ERK1/2 with the actin cytoskeleton regulates the outcome of ERK1/2 signaling.
authors.affiliation: Department of Health Sciences, Boston University, Boston, Massachusetts, United States of America. svetter@bu.edu
authors.forename: Susanne
authors.initials: S
authors.lastname: Vetterkind
authors.forename: Robert J
authors.initials: RJ
authors.lastname: Saphirstein
authors.forename: Kathleen G
authors.initials: KG
authors.lastname: Morgan
mesh_terms: D008841:Actin Cytoskeleton / Q000378:metabolism; D000199:Actins / Q000378:metabolism*; D000818:Animals; D002148:Calmodulin-Binding Proteins / Q000378:metabolism; D051242:Caveolin 1 / Q000378:metabolism; D002460:Cell Line; D004789:Enzyme Activation / Q000187:drug effects; D055785:Gene Knockdown Techniques; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism*; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism*; D008954:Models, Biological; D032389:Myocytes, Smooth Muscle / Q000187:drug effects / Q000201:enzymology*; D010766:Phosphorylation / Q000187:drug effects; D021381:Protein Transport / Q000187:drug effects; D051381:Rats; D044967:Serum; D012995:Solubility / Q000187:drug effects; D013347:Subcellular Fractions / Q000187:drug effects / Q000201:enzymology; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
chemical_list: D000199:Actins; D002148:Calmodulin-Binding Proteins; D051242:Caveolin 1; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D013755:Tetradecanoylphorbol Acetate