Role of myosin II activity and the regulation of myosin light chain phosphorylation in astrocytomas.
The generation of contractile force mediated by actin-myosin interactions is essential for cell motility. Myosin activity is promoted by phosphorylation of myosin light chain (MLC). MLC phosphorylation in large part is controlled by kinases that are effectors of Rho family GTPases. Accordingly, in this study we examined the effects of ROCK and Rac1 inhibition on MLC phosphorylation in astrocytoma cells. We found that low concentrations of the ROCK inhibitor Y27632 increased the phosphorylation state of the Triton X-100 soluble fraction of MLC, whereas higher concentrations of Y27632 decreased soluble phospho-MLC. These effects of Y27632 were dependent on Rac1. The soluble form of phospho-MLC comprises about 10% of total phospho-MLC in control cells. Interestingly, ROCK inhibition led to a decrease in the phosphorylation state of total MLC, whereas Rac1 inhibition had little effect. Thus, the soluble form of MLC is differentially regulated by ROCK and Rac1 compared with MLC examined in a total cell extract. We also observed that astrocytoma migration is stimulated by low concentrations of the myosin II inhibitor blebbistatin. However, higher concentrations of blebbistatin inhibit migration leading us to believe that migration has a biphasic dependence on myosin II activity. Taken together, our data show that modulation of myosin II activity is important in determining optimal astrocytoma migration. In addition, these findings suggest that there are at least two populations of MLC that are differentially regulated.
Salhia
Bodour
B
The Arthur and Sonia Labatt Brain Tumor Research Center, The Hospital for Sick Children, The University of Toronto, Toronto, Ontario, Canada.
Hwang
Jeong Hyun
JH
Smith
Christian A
CA
Nakada
Mitsutoshi
M
Rutka
Fiona
F
Symons
Marc
M
Rutka
James T
JT
D001254:Astrocytoma / Q000378:metabolism* / Q000473:pathology; D045744:Cell Line, Tumor; D002465:Cell Movement / Q000502:physiology; D006801:Humans; D018994:Myosin Light Chains / Q000378:metabolism*; D024462:Myosin Type II / Q000378:metabolism / Q000502:physiology*; D010766:Phosphorylation; D020830:rac1 GTP-Binding Protein / Q000502:physiology; D054460:rho-Associated Kinases / Q000502:physiology
D018994:Myosin Light Chains; C107953:RAC1 protein, human; D054460:rho-Associated Kinases; D024462:Myosin Type II; D020830:rac1 GTP-Binding Protein