N-WASP involvement in dorsal ruffle formation in mouse embryonic fibroblasts

Legg, J. A., Bompard, G., Dawson, J., Morris, H. L., Andrew, N., Cooper, L., Johnston, S. A., Tramountanis, G. and Machesky, L. (2007) N-WASP involvement in dorsal ruffle formation in mouse embryonic fibroblasts. Molecular Biology of the Cell, 18(2), pp. 678-687. (doi: 10.1091/mbc.e06-06-0569) (PMID:17182853) (PMCID:PMC1783773)

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Abstract

The Wiskott–Aldrich syndrome protein (WASP) family activates the Arp2/3 complex leading to the formation of new actin filaments. Here, we study the involvement of Scar1, Scar2, N-WASP, and Arp2/3 complex in dorsal ruffle formation in mouse embryonic fibroblasts (MEFs). Using platelet-derived growth factor to stimulate circular dorsal ruffle assembly in primary E13 and immortalized E9 Scar1+/+ and Scar1 null MEFs, we establish that Scar1 loss does not impair the formation of dorsal ruffles. Reduction of Scar2 protein levels via small interfering RNA (siRNA) also did not affect dorsal ruffle production. In contrast, wiskostatin, a chemical inhibitor of N-WASP, potently suppressed dorsal ruffle formation in a dose-dependent manner. Furthermore, N-WASP and Arp2 siRNA treatment significantly decreased the formation of dorsal ruffles in MEFs. In addition, the expression of an N-WASP truncation mutant that cannot bind Arp2/3 complex blocked the formation of these structures. Finally, N-WASP−/− fibroblast-like cells generated aberrant dorsal ruffles. These ruffles were highly unstable, severely depleted of Arp2/3 complex, and diminished in size. We hypothesize that N-WASP and Arp2/3 complex are part of a multiprotein assembly important for the generation of dorsal ruffles and that Scar1 and Scar2 are dispensable for this process.

Item Type:Articles
Additional Information:J.A.L. and S.A.J. were supported by Biotechnology and Biological Sciences Research Council studentships, L.M.M. is supported by a Medical Research Council Senior Research Fellowship G117/569, and G.B. and J.D. were supported by a project grant to L.M.M. from the Association for International Cancer Research. G.T. was supported by a European 5th Framework grant. L.C. was supported by a Human Frontier Science Program Organization grant.
Status:Published
Refereed:Yes
Glasgow Author(s) Enlighten ID:Machesky, Professor Laura
Authors: Legg, J. A., Bompard, G., Dawson, J., Morris, H. L., Andrew, N., Cooper, L., Johnston, S. A., Tramountanis, G., and Machesky, L.
College/School:College of Medical Veterinary and Life Sciences > Institute of Cancer Sciences
Journal Name:Molecular Biology of the Cell
Publisher:American Society for Cell Biology
ISSN:1939-4586
ISSN (Online):1939-4586
Published Online:20 December 2006

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