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© The Rockefeller University Press,
0021-9525/2000//697 $5.00
The Journal of Cell Biology, Volume 151, Number 3,
, 2000 697-708
Original Article |
Localization and Activity of Myosin Light Chain Kinase Isoforms during the Cell Cycle
Phosphorylation on Ser 19 of the myosin II regulatory light chain by myosin light chain kinase (MLCK) regulates actomyosin contractility in smooth muscle and vertebrate nonmuscle cells. The smooth/nonmuscle MLCK gene locus produces two kinases, a high molecular weight isoform (long MLCK) and a low molecular weight isoform (short MLCK), that are differentially expressed in smooth and nonmuscle tissues. To study the relative localization of the MLCK isoforms in cultured nonmuscle cells and to determine the spatial and temporal dynamics of MLCK localization during mitosis, we constructed green fluorescent protein fusions of the long and short MLCKs. In interphase cells, localization of the long MLCK to stress fibers is mediated by five DXRXXL motifs, which span the junction of the NH2-terminal extension and the short MLCK. In contrast, localization of the long MLCK to the cleavage furrow in dividing cells requires the five DXRXXL motifs as well as additional amino acid sequences present in the NH2-terminal extension. Thus, it appears that nonmuscle cells utilize different mechanisms for targeting the long MLCK to actomyosin structures during interphase and mitosis. Further studies have shown that the long MLCK has twofold lower kinase activity in early mitosis than in interphase or in the early stages of postmitotic spreading. These findings suggest a model in which MLCK and the myosin II phosphatase (Totsukawa, G., Y. Yamakita, S. Yamashiro, H. Hosoya, D.J. Hartshorne, and F. Matsumura. 1999. J. Cell Biol. 144:735–744) act cooperatively to regulate the level of Ser 19–phosphorylated myosin II during mitosis and initiate cytokinesis through the activation of myosin II motor activity.
Key Words: myosin • phosphorylation • myosin light chain kinase • cell division • cytokinesis
© 2000 The Rockefeller University Press
A. Poperechnaya and O. Varlamova contributed equally to this work.Abbreviations used in this paper: CaM kinase II, Ca2+/calmodulin-dependent protein kinase; GFP, green fluorescent protein; HMM, heavy meromyosin; MBS, myosin binding subunit; MLCK, myosin light chain kinase; PAK, p21-activated kinase; PKA, cAMP-dependent protein kinase; PKC, protein kinase C; RLC, regulatory light chain.
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