Regulation of podosome dynamics by WASp phosphorylation: implication in matrix degradation and chemotaxis in macrophages.
Podosomes, adhesion structures capable of matrix degradation, have been linked with the ability of cells to perform chemotaxis and invade tissues. Wiskott-Aldrich Syndrome protein (WASp), an effector of the RhoGTPase Cdc42 and a Src family kinase substrate, regulates macrophage podosome formation. In this study, we demonstrate that WASp is active in podosomes by using TIRF-FRET microscopy. Pharmacological and RNA interference approaches suggested that continuous WASp activity is required for podosome formation and function. Rescue experiments using point mutations demonstrate an absolute requirement for Cdc42 binding to WASp in podosome formation. Although tyrosine phosphorylation was not absolutely required for podosome formation, phosphorylation did regulate the rate of podosome nucleation and actin filament stability. Importantly, WASp tyrosine phosphorylation does not alter WASp activation, instead phosphorylation appears to be important for the restriction of WASp activity to podosomes. In addition, the matrix-degrading ability of cells requires WASp phosphorylation. Chemotactic responses to CSF-1 were also attenuated in the absence of endogenous WASp, which could not be rescued with either tyrosine mutation. These results suggest a more complex role for tyrosine phosphorylation than simply in the regulation of WASp activity, and suggest a link between podosome dynamics and macrophage migration.
Dovas
Athanassios
A
Department of Anatomy and Structural Biology, Albert Einstein College of Medicine, Bronx, NY 10461, USA.
Gevrey
Jean-Claude
JC
Grossi
Alberto
A
Park
Haein
H
Abou-Kheir
Wassim
W
Cox
Dianne
D
D000818:Animals; D002460:Cell Line; D022081:Cell Surface Extensions / Q000235:genetics / Q000378:metabolism*; D002478:Cells, Cultured; D002633:Chemotaxis; D005109:Extracellular Matrix / Q000378:metabolism*; D006801:Humans; D008264:Macrophages / Q000502:physiology*; D010766:Phosphorylation; D011485:Protein Binding; D051316:Wiskott-Aldrich Syndrome Protein / Q000235:genetics / Q000378:metabolism*; D020764:cdc42 GTP-Binding Protein / Q000235:genetics / Q000378:metabolism
D051316:Wiskott-Aldrich Syndrome Protein; D020764:cdc42 GTP-Binding Protein