Department of Cell Biology; Department of Physiology
Actins; Animals; COS Cells; Cell Adhesion; Cell Movement; Cercopithecus aethiops; Humans; Isoenzymes; Membrane Proteins; Mice; Microfilament Proteins; Myosin Type II; Myosin-Light-Chain Kinase; RNA Interference; Recombinant Fusion Proteins
Cell Biology | Life Sciences | Medicine and Health Sciences
During cell migration, myosin II modulates adhesion, cell protrusion and actin organization at the leading edge. We show that an F-actin- and membrane-associated scaffolding protein, called supervillin (SV, p205), binds directly to the subfragment 2 domains of nonmuscle myosin IIA and myosin IIB and to the N-terminus of the long form of myosin light chain kinase (L-MLCK). SV inhibits cell spreading via an MLCK- and myosin II-dependent mechanism. Overexpression of SV reduces the rate of cell spreading, and RNAi-mediated knockdown of endogenous SV increases it. Endogenous and EGFP-tagged SV colocalize with, and enhance the formation of, cortical bundles of F-actin and activated myosin II during early cell spreading. The effects of SV are reversed by inhibition of myosin heavy chain (MHC) ATPase (blebbistatin), MLCK (ML-7) or MEK (U0126), but not by inhibiting Rho-kinase with Y-27632. Flag-tagged L-MLCK co-localizes in cortical bundles with EGFP-SV, and kinase-dead L-MLCK disorganizes these bundles. The L-MLCK- and myosin-binding site in SV, SV1-171, rearranges and co-localizes with mono- and di-phosphorylated myosin light chain and with L-MLCK, but not with the short form of MLCK (S-MLCK) or with myosin phosphatase. Thus, the membrane protein SV apparently contributes to myosin II assembly during cell spreading by modulating myosin II regulation by L-MLCK.
DOI of Published Version
J Cell Sci. 2007 Nov 1;120(Pt 21):3792-803. Epub 2007 Oct 9. Link to article on publisher's site
Journal of cell science
Takizawa, Norio; Ikebe, Reiko; Ikebe, Mitsuo; and Luna, Elizabeth J., "Supervillin slows cell spreading by facilitating myosin II activation at the cell periphery" (2007). Open Access Articles. 1283.