The goal of this proposal is to elucidate the molecular mechanisms of cell division and cell migration, by defining the specific roles of phosphorylation of the regulatory light chain of myosin II (MLC) in these processes. Our studies focus on a critical pathway controlled by myosin phosphatase targeting subunit (MYPT), and myosin light chain kinase (MLCK).
Specific aim 1 is to determine the function(s) of MYPT in spindle assembly and cytokinesis. MYPT is phosphorylated by cdc2 kinase, resulting in complex formation with polo-like kinase 1 (PLK1). MYPT depletion delays the prometaphase/metaphase transition and causes cytokinesis failure. Because PLK1 is an essential kinase for spindle assembly, as well as cytokinesis, we hypothesize that the MYPT-PLK1 association regulates these processes.
Specific aim 2 is to determine the roles of MLCK and MYPT in directed cell migration, and to identify the upstream molecules that control MLCK and MYPT. MLCK controls MLC phosphorylation at the cell periphery and restricts membrane protrusions during cell migration. Phospho-specific antibodies against MLCK and MYPT, as well as phosphorylation-site mutants will be generated to determine which upstream kinases provide critical spatial regulation to MLCK and MYPT.
Specific aim 3 is to determine the mechanism by which MLC phosphorylation controls the assembly of cortical actin and regulates cell migration in a 3D matrix. In the presence of protease inhibitors, a tumor cell adapts its cell shape and mode of migration to overcome the physical barrier of the matrix: it constricts its cell body and moves like an amoeba. Analyses will be performed to determine when and where MLC phosphorylation occurs during constriction-mediated amoeboid movement of tumor cells, and to identify which upstream molecules are essential for the assembly and function of the constriction ring. These studies will provide new insights, not only into how normal cells divide and move, but also into two fundamental aspects of cancer pathophysiology, the loss of cell division control and metastasis.
| Matsumura, Fumio; Yamakita, Yoshihiko; Yamashiro, Shigeko (2011) Myosin light chain kinases and phosphatase in mitosis and cytokinesis. Arch Biochem Biophys 510:76-82 |
| Matsumura, Fumio; Yamakita, Yoshihiko; Yamashiro, Shigeko (2011) Myosin phosphatase-targeting subunit 1 controls chromatid segregation. J Biol Chem 286:10825-33 |
| Yamakita, Yoshihiko; Matsumura, Fumio; Lipscomb, Michael W et al. (2011) Fascin1 promotes cell migration of mature dendritic cells. J Immunol 186:2850-9 |
| Yamakita, Yoshihiko; Matsumura, Fumio; Yamashiro, Shigeko (2009) Fascin1 is dispensable for mouse development but is favorable for neonatal survival. Cell Motil Cytoskeleton 66:524-34 |
| Yamashiro, Shigeko; Yamakita, Yoshihiko; Totsukawa, Go et al. (2008) Myosin phosphatase-targeting subunit 1 regulates mitosis by antagonizing polo-like kinase 1. Dev Cell 14:787-97 |
| Matsumura, Fumio; Hartshorne, David J (2008) Myosin phosphatase target subunit: Many roles in cell function. Biochem Biophys Res Commun 369:149-56 |
| Alemi, Mansour; Prigione, Alessandro; Wong, Alice et al. (2007) Mitochondrial DNA deletions inhibit proteasomal activity and stimulate an autophagic transcript. Free Radic Biol Med 42:32-43 |
| Lu, Chunye; Cortopassi, Gino (2007) Frataxin knockdown causes loss of cytoplasmic iron-sulfur cluster functions, redox alterations and induction of heme transcripts. Arch Biochem Biophys 457:111-22 |
| Napoli, Eleonora; Taroni, Franco; Cortopassi, Gino A (2006) Frataxin, iron-sulfur clusters, heme, ROS, and aging. Antioxid Redox Signal 8:506-16 |
| Takiguchi, Kingo; Matsumura, Fumio (2005) Role of the basic C-terminal half of caldesmon in its regulation of F-actin: comparison between caldesmon and calponin. J Biochem 138:805-13 |
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