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Am J Physiol Cell Physiol 284: C627-C639, 2003. First published October 30, 2002; doi:10.1152/ajpcell.00137.2002
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Vol. 284, Issue 3, C627-C639, March 2003

External mechanical strain regulates membrane targeting of Rho GTPases by controlling microtubule assembly

Andrew J. Putnam1, James J. Cunningham1, Brendan B. L. Pillemer2, and David J. Mooney1,3,4

Departments of 1 Chemical Engineering, 2 Microbiology, 3 Biomedical Engineering, and 4 Biologic and Materials Sciences, University of Michigan, Ann Arbor, Michigan 48109-1078

Transmission of externally applied mechanical forces to the interior of a cell requires coordination of biochemical signaling pathways with changes in cytoskeletal assembly and organization. In this study, we addressed one potential mechanism for this signal integration by applying uniform single external mechanical strains to aortic smooth muscle cells (SMCs) via their adhesion substrate. A tensile strain applied to the substrate for 15 min significantly increased microtubule (MT) assembly by 32 ± 7%, with no apparent effect on the cells' focal adhesions as revealed by immunofluorescence and quantitative analysis of Triton X-100-insoluble vinculin levels. A compressive strain decreased MT mass by 24 ± 9% but did not influence the level of vinculin in focal adhesions. To understand the decoupling of these two cell responses to mechanical strain, we examined a redistribution of the small GTPases RhoA and Rac. Tensile strain was found to decrease the amount of membrane-associated RhoA and Rac by 70 ± 9% and 45 ± 11%, respectively, compared with static controls. In contrast, compressive strain increased membrane-associated RhoA and Rac levels by 74 ± 17% and 36 ± 13%, respectively. Disruption of the MT network by prolonged treatments with low doses of either nocodazole or paclitaxel before the application of strain abolished the redistribution of RhoA and Rac in response to the applied forces. Combined, these results indicate that the effects of externally applied mechanical strain on the distribution and activation of the Rho family GTPases require changes in the state of MT polymerization.

cytoskeleton; mechanotransduction; focal adhesions


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