title: Myosin light chain kinase/actin interaction in phorbol dibutyrate-stimulated smooth muscle cells.
abstract: Previous work has suggested that in addition to its kinase activity, myosin light chain kinase (MLCK) exhibits non-kinase properties within its N-terminus that could influence cytoskeletal organization of smooth muscle cells (A. Nakamura et al. Biochem Biophys Res Commun. 2008;369:135-143). Myosin ATPase activity measurements indicate that the 26-41 peptide of MLCK significantly decreases ATPase activity as the concentration of this peptide increases. Sliding velocity of actin-filaments on myosin and stress responses in skinned smooth muscle tissue are also inhibited. Peptide-mediated uptake and the microinjection technique in cells indicate that the peptide was necessary for actin-filament stabilization. Fluorescence resonance energy transfer analysis indicated that in the presence of MLCK, α-actin but not β-actin remodeled during phorbol 12,13-dibutyrate (PDBu)-induced contractions. PDBu also induced podosomes in the cell. When MLCK expression was down-regulated by introduction of RNAi for MLCK by lentivirus vector into the cells, we failed to observe the podosome induction upon PDBu stimulation. Rescue experiments indicate that the non-kinase activity of MLCK plays an important role in maintaining actin stress fibers and in the PDBu-induced reorganization of actin-filaments in smooth muscle cells.
authors.affiliation: Department of Pharmacology, Physiology, and Toxicology, The Joan C. Edwards School of Medicine, Marshall University, Huntington, WV 25704, USA.
authors.forename: Sean E
authors.initials: SE
authors.lastname: Thatcher
authors.forename: Mike E
authors.initials: ME
authors.lastname: Fultz
authors.forename: Hideyuki
authors.initials: H
authors.lastname: Tanaka
authors.forename: Haruo
authors.initials: H
authors.lastname: Hagiwara
authors.forename: Hou-Li
authors.initials: HL
authors.lastname: Zhang
authors.forename: Ying
authors.initials: Y
authors.lastname: Zhang
authors.forename: Kohichi
authors.initials: K
authors.lastname: Hayakawa
authors.forename: Shinji
authors.initials: S
authors.lastname: Yoshiyama
authors.forename: Akio
authors.initials: A
authors.lastname: Nakamura
authors.forename: Hong Hui
authors.initials: HH
authors.lastname: Wang
authors.forename: Takeshi
authors.initials: T
authors.lastname: Katayama
authors.forename: Masaru
authors.initials: M
authors.lastname: Watanabe
authors.forename: Yuan
authors.initials: Y
authors.lastname: Lin
authors.forename: Gary L
authors.initials: GL
authors.lastname: Wright
authors.forename: Kazuhiro
authors.initials: K
authors.lastname: Kohama
mesh_terms: D000199:Actins / Q000378:metabolism*; D000818:Animals; D002460:Cell Line; D022081:Cell Surface Extensions / Q000187:drug effects / Q000378:metabolism / Q000648:ultrastructure; D002645:Chickens; D003599:Cytoskeleton / Q000187:drug effects / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000378:metabolism; D020868:Gene Silencing; D006168:Guinea Pigs; D066298:In Vitro Techniques; D007700:Kinetics; D008840:Microfilament Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D009119:Muscle Contraction / Q000187:drug effects*; D009130:Muscle, Smooth / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure; D032389:Myocytes, Smooth Muscle / Q000187:drug effects / Q000378:metabolism / Q000648:ultrastructure; D009219:Myosin-Light-Chain Kinase / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D009218:Myosins / Q000037:antagonists & inhibitors; D010446:Peptide Fragments / Q000378:metabolism; D015240:Phorbol 12,13-Dibutyrate / Q000494:pharmacology*; D010766:Phosphorylation / Q000187:drug effects; D020033:Protein Isoforms / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D034741:RNA, Small Interfering; D051381:Rats
chemical_list: D000199:Actins; D004791:Enzyme Inhibitors; D008840:Microfilament Proteins; D010446:Peptide Fragments; D020033:Protein Isoforms; D034741:RNA, Small Interfering; D015240:Phorbol 12,13-Dibutyrate; D009219:Myosin-Light-Chain Kinase; D009218:Myosins