Myosin light chain kinase and acto-myosin contractility modulate activation of the ERK cascade downstream of oncogenic Ras.
The actin cytoskeleton is recognized as an important component of both adhesion- and growth factor-dependent signaling, but its role in oncogene-dependent signaling has received much less attention. In this study, we investigated the role played by the acto-myosin cytoskeleton and its main regulators, i.e., myosin light chain kinase and Rho kinase, in oncogenic Ki-Ras-induced signaling. We found that activation of the ERK cascade by Ras is dependent on acto-myosin contractility, under the regulation of myosin light chain kinase but not Rho kinase. Inhibition of myosin II or myosin light chain kinase caused a complete loss of ERK phosphorylation in a time- and dose-dependent manner, but proved dispensable for activation of the PI3K pathway. We also provide evidence that the target of myosin light chain kinase lays at the level of Raf activation. Since myosin light chain kinase is a target of ERK, these results suggest a previously uncharacterized signaling pathway involving Ras-mediated alterations of the actin cytoskeleton, which might play a critical role in ERK activation by the Ras oncogene and contribute to aberrant signaling and enhanced cell motility. In addition, restoration of stress fibers following ectopic expression of tropomyosin 2 resulted in reduced levels of ERK phosphorylation. Finally, these studies suggest that myosin light chain kinase but not Rho kinase plays an essential role in the generation of ERK signaling in transformed cells and indicate distinct cellular roles for Rho-kinase and myosin light chain kinase-dependent functions involving the regulation of acto-myosin contractility.
Helfman
David M
DM
Department of Cell Biology and Anatomy, University of Miami School of Medicine, Sylvester Comprehensive Cancer Center, Miami, Florida 33136, USA. dhelfman@med.miami.edu
Pawlak
Geraldine
G
D000205:Actomyosin / Q000378:metabolism*; D000818:Animals; D002448:Cell Adhesion; D002465:Cell Movement; D004789:Enzyme Activation; D005455:Fluorescent Antibody Technique; D011905:Genes, ras / Q000502:physiology*; D015151:Immunoblotting; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism*; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism*; D009119:Muscle Contraction; D018482:Muscle, Skeletal / Q000166:cytology / Q000378:metabolism; D009219:Myosin-Light-Chain Kinase / Q000378:metabolism*; D010766:Phosphorylation; D019908:Proto-Oncogene Proteins c-raf / Q000378:metabolism; D051381:Rats; D012190:Retroviridae; D015398:Signal Transduction; D020741:rho GTP-Binding Proteins / Q000378:metabolism
D000205:Actomyosin; D019908:Proto-Oncogene Proteins c-raf; D009219:Myosin-Light-Chain Kinase; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020741:rho GTP-Binding Proteins