Myosin Light Chain Kinase / Actin Interaction in Phorbol Dibutyrate-Stimulated Smooth Muscle Cells

• Main Authors: Sean E. Thatcher (Author), Mike E. Fultz (Author), Hideyuki Tanaka (Author), Haruo Hagiwara (Author), Hou-Li Zhang (Author), Ying Zhang (Author), Kohichi Hayakawa (Author), Shinji Yoshiyama (Author), Akio Nakamura (Author), Hong Hui Wang (Author), Takeshi Katayama (Author), Masaru Watanabe (Author), Yuan Lin (Author), Gary L. Wright (Author), Kazuhiro Kohama (Author)
• Format: Book
• Published: Elsevier, 2011-01-01T00:00:00Z.
• Subjects: article
• Online Access: Connect to this object online.

Summary

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. Keywords:: actin-remodeling, podosome, phorbol 12,13-dibutyrate (PDBu), FRET analysis, myosin light chain kinase (MLCK)-deficient cell

Item Description

1347-8613
10.1254/jphs.10296FP