CYTOSKELETAL REMODELING Ruffling with Rab5 and RN-tre

Lanzetti et al. investigated receptor tyrosine kinase (RTK)-mediated regulation of the actin cytoskeleton and discovered that Rab5, a small guanosine triphosphatase (GTPase) previously implicated in intracellular trafficking, is critical to circular ruffling (an RTK-dependent process that involves cytoskeletal remodeling). Expression of a Rab5 dominant-negative mutant in mouse embryo fibroblasts (MEFs) inhibited platelet-derived growth factor (PDGF)-mediated production of circular ruffles, without affecting edge ruffles (which depend on Ras), as did overexpression of the Rab5-specific GTPase-activating protein (GAP) RN-tre. Experiments in which MEFs were transfected with wild-type or mutant forms of Rab5, Ras, Rac, or phosphatidylinositol 3-kinase (PI3K) alone or in combination suggested that independent signals from Rab5, PI3K, and Rac were all required for circular ruffling. Mutational analysis indicated that suppression of circular ruffling (and the related process of macropinocytosis) by RN-tre did not depend solely on its GAP activity but also on a region in its C terminus: Only a mutant that lacked both GAP activity and this region failed to inhibit PDGF-dependent circular ruffling. The authors used mass spectrometry to show that actinin-4 (ACTN4), a nonmuscle isoform of -actinin, interacted with this region of RN-tre. Endogenous RN-tre and ACTN4 exhibited PDGF-dependent coimmunoprecipitation, and PDGF-dependent ruffling was impaired in cells in which RN-tre or ACTN4 expression was inhibited through RNA interference. RN-tre interacted with F-actin through regions distinct from the region that bound ACTN4. Thus, Rab5 mediates RTK-dependent cytoskeletal signals involved in circular ruffling and RN-tre acts as a Rab5 effector--possibly by connecting Rab5, ACTN4 and F-actin--as well as a Rab5 GAP.

L. Lanzetti, A. Palamidessi, L. Areces, G. Scita, P. P. Di Fiore, Rab5 is a signalling GTPase involved in actin remodelling by receptor tyrosine kinases. Nature 429, 309-314 (2004). [Online Journal]