Traction Forces of Fibroblasts Are Regulated by the Rho-dependent Kinase but Not by the Myosin Light Chain Kinase

Overview

Journal Arch Biochem Biophys

Publisher Elsevier

Specialties Biochemistry
Biophysics

Date 2006 Nov 11

PMID 17094935

Citations 66

Authors

Karen A Beningo

Kozue Hamao

Micah Dembo

Yu-Li Wang

Hiroshi Hosoya

Affiliations

Soon will be listed here.

Abstract

Adhesive cells show complex mechanical interactions with the substrate, however the exact mechanism of such interactions, termed traction forces, is still unclear. To address this question we have measured traction forces of fibroblasts treated with agents that affect the myosin II-dependent contractile mechanism. Using the potent myosin II inhibitor blebbistatin, we demonstrate that traction forces are strongly dependent on a functional myosin II heavy chain. Since myosin II is regulated by both the myosin light chain kinase (MLCK) and, directly or indirectly, the Rho-associated kinase (ROCK), we examined the effects of inhibitors against these kinases. Interestingly, inhibition of the myosin light chain kinase had no detectable effect, while inhibition of the Rho-dependent kinase caused strong inhibition of traction forces. Our results indicate that ROCK and MLCK play non-redundant roles in regulating myosin II functions, and that a subset of myosin II, regulated by the Rho small GTPase, may be responsible for the regulation of traction forces in migrating fibroblasts.

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References

Smilenov L, Mikhailov A, Pelham R, Marcantonio E, Gundersen G . Focal adhesion motility revealed in stationary fibroblasts. Science. 1999; 286(5442):1172-4. DOI: 10.1126/science.286.5442.1172. View

Abe M, Ho C, Kamm K, Grinnell F . Different molecular motors mediate platelet-derived growth factor and lysophosphatidic acid-stimulated floating collagen matrix contraction. J Biol Chem. 2003; 278(48):47707-12. DOI: 10.1074/jbc.M306228200. View

Totsukawa G, Yamakita Y, Yamashiro S, Hartshorne D, Sasaki Y, Matsumura F . Distinct roles of ROCK (Rho-kinase) and MLCK in spatial regulation of MLC phosphorylation for assembly of stress fibers and focal adhesions in 3T3 fibroblasts. J Cell Biol. 2000; 150(4):797-806. PMC: 2175273. DOI: 10.1083/jcb.150.4.797. View

Amano M, Fukata Y, Kaibuchi K . Regulation and functions of Rho-associated kinase. Exp Cell Res. 2000; 261(1):44-51. DOI: 10.1006/excr.2000.5046. View

Munevar S, Wang Y, Dembo M . Traction force microscopy of migrating normal and H-ras transformed 3T3 fibroblasts. Biophys J. 2001; 80(4):1744-57. PMC: 1301364. DOI: 10.1016/s0006-3495(01)76145-0. View

Katoh K, Kano Y, Amano M, Kaibuchi K, Fujiwara K . Stress fiber organization regulated by MLCK and Rho-kinase in cultured human fibroblasts. Am J Physiol Cell Physiol. 2001; 280(6):C1669-79. DOI: 10.1152/ajpcell.2001.280.6.C1669. View

Beningo K, Dembo M, Kaverina I, Small J, Wang Y . Nascent focal adhesions are responsible for the generation of strong propulsive forces in migrating fibroblasts. J Cell Biol. 2001; 153(4):881-8. PMC: 2192381. DOI: 10.1083/jcb.153.4.881. View

Jin Y, Blue E, Dixon S, Hou L, Wysolmerski R, Gallagher P . Identification of a new form of death-associated protein kinase that promotes cell survival. J Biol Chem. 2001; 276(43):39667-78. PMC: 2823794. DOI: 10.1074/jbc.M101886200. View

Saitoh T, Takemura S, Ueda K, Hosoya H, Nagayama M, Haga H . Differential localization of non-muscle myosin II isoforms and phosphorylated regulatory light chains in human MRC-5 fibroblasts. FEBS Lett. 2001; 509(3):365-9. DOI: 10.1016/s0014-5793(01)03186-6. View

10.

Fukuta Y, Ueda K, Iwasaki T, Hosoya H . HeLa ZIP kinase induces diphosphorylation of myosin II regulatory light chain and reorganization of actin filaments in nonmuscle cells. Oncogene. 2002; 20(57):8175-83. DOI: 10.1038/sj.onc.1205055. View

11.

Chew T, Wolf W, Gallagher P, Matsumura F, Chisholm R . A fluorescent resonant energy transfer-based biosensor reveals transient and regional myosin light chain kinase activation in lamella and cleavage furrows. J Cell Biol. 2002; 156(3):543-53. PMC: 2173328. DOI: 10.1083/jcb.200110161. View

12.

Beningo K, Wang Y . Flexible substrata for the detection of cellular traction forces. Trends Cell Biol. 2002; 12(2):79-84. DOI: 10.1016/s0962-8924(01)02205-x. View

13.

Beningo K, Lo C, Wang Y . Flexible polyacrylamide substrata for the analysis of mechanical interactions at cell-substratum adhesions. Methods Cell Biol. 2002; 69:325-39. DOI: 10.1016/s0091-679x(02)69021-1. View

14.

Uchimura T, Fumoto K, Yamamoto Y, Ueda K, Hosoya H . Spatial localization of mono-and diphosphorylated myosin II regulatory light chain at the leading edge of motile HeLa cells. Cell Struct Funct. 2003; 27(6):479-86. DOI: 10.1247/csf.27.479. View

15.

Fumoto K, Uchimura T, Iwasaki T, Ueda K, Hosoya H . Phosphorylation of myosin II regulatory light chain is necessary for migration of HeLa cells but not for localization of myosin II at the leading edge. Biochem J. 2002; 370(Pt 2):551-6. PMC: 1223179. DOI: 10.1042/BJ20021559. View

16.

Straight A, Cheung A, Limouze J, Chen I, Westwood N, Sellers J . Dissecting temporal and spatial control of cytokinesis with a myosin II Inhibitor. Science. 2003; 299(5613):1743-7. DOI: 10.1126/science.1081412. View

17.

Somlyo A, Somlyo A . Ca2+ sensitivity of smooth muscle and nonmuscle myosin II: modulated by G proteins, kinases, and myosin phosphatase. Physiol Rev. 2003; 83(4):1325-58. DOI: 10.1152/physrev.00023.2003. View

18.

Bershadsky A, Balaban N, Geiger B . Adhesion-dependent cell mechanosensitivity. Annu Rev Cell Dev Biol. 2003; 19:677-95. DOI: 10.1146/annurev.cellbio.19.111301.153011. View

19.

Ridley A, Schwartz M, Burridge K, Firtel R, Ginsberg M, Borisy G . Cell migration: integrating signals from front to back. Science. 2003; 302(5651):1704-9. DOI: 10.1126/science.1092053. View

20.

Totsukawa G, Wu Y, Sasaki Y, Hartshorne D, Yamakita Y, Yamashiro S . Distinct roles of MLCK and ROCK in the regulation of membrane protrusions and focal adhesion dynamics during cell migration of fibroblasts. J Cell Biol. 2004; 164(3):427-39. PMC: 2172229. DOI: 10.1083/jcb.200306172. View