MYBPH inhibits NM IIA assembly via direct interaction with NMHC IIA and reduces cell motility.
Actomyosin filament assembly is a critical step in tumor cell migration. We previously found that myosin binding protein H (MYBPH) is directly transactivated by the TTF-1 lineage-survival oncogene in lung adenocarcinomas and inhibits phosphorylation of the myosin regulatory light chain (RLC) of non-muscle myosin IIA (NM IIA) via direct interaction with Rho kinase 1 (ROCK1). Here, we report that MYBPH also directly interacts with an additional molecule, non-muscle myosin heavy chain IIA (NMHC IIA), which was found to occur between MYBPH and the rod portion of NMHC IIA. MYBPH inhibited NMHC IIA assembly and reduced cell motility. Conversely, siMYBPH-induced increased motility was partially, yet significantly, suppressed by blebbistatin, a non-muscle myosin II inhibitor, while more profound effects were attained by combined treatment with siROCK1 and blebbistatin. Electron microscopy observations showed well-ordered paracrystals of NMHC IIA reflecting an assembled state, which were significantly less frequently observed in the presence of MYBPH. Furthermore, an in vitro sedimentation assay showed that a greater amount of NMHC IIA was in an unassembled state in the presence of MYBPH. Interestingly, treatment with a ROCK inhibitor that impairs transition of NM IIA from an assembly-incompetent to assembly-competent state reduced the interaction between MYBPH and NMHC IIA, suggesting that MYBPH has higher affinity to assembly-competent NM IIA. These results suggest that MYBPH inhibits RLC and NMHC IIA, independent components of NM IIA, and negatively regulates actomyosin organization at 2 distinct steps, resulting in firm inhibition of NM IIA assembly.
Division of Molecular Carcinogenesis, Center for Neurological Diseases and Cancer, Nagoya University Graduate School of Medicine, Nagoya 466-8550, Japan.
Yasuyuki
Y
Hosono
Jiro
J
Usukura
Tomoya
T
Yamaguchi
Kiyoshi
K
Yanagisawa
Motoshi
M
Suzuki
Takashi
T
Takahashi
D000205:Actomyosin / Q000378:metabolism; D000230:Adenocarcinoma / Q000378:metabolism; D000077192:Adenocarcinoma of Lung; D045744:Cell Line, Tumor; D002465:Cell Movement; D003598:Cytoskeletal Proteins / Q000378:metabolism*; D006801:Humans; D008175:Lung Neoplasms / Q000378:metabolism; D020409:Molecular Motor Proteins / Q000378:metabolism*; D018995:Myosin Heavy Chains / Q000378:metabolism*; D024507:Nonmuscle Myosin Type IIA / Q000037:antagonists & inhibitors* / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics; D054460:rho-Associated Kinases / Q000037:antagonists & inhibitors / Q000235:genetics
D003598:Cytoskeletal Proteins; C484119:MYBPH protein, human; C103814:MYH9 protein, human; D020409:Molecular Motor Proteins; D034741:RNA, Small Interfering; D000205:Actomyosin; C515851:ROCK1 protein, human; D054460:rho-Associated Kinases; D024507:Nonmuscle Myosin Type IIA; D018995:Myosin Heavy Chains