Rab5 expression (red) into HeLa cells promotes the formation of actin-rich (green) dorsal circular ruffles in 2D (left) and the extension of multiple elongated protrusions in 3D matrices (right).

Image courtesy of Dr Giorgio Scita, IFOM, The FIRC Institute of Molecular Oncology Foundation, Milan, Italy.

Growth factor signalling is mediated by membrane receptor tyrosine kinases (RTKs) and induces cell motility via the activation of Rac — a small Rho GTPase known to be a key coordinator in actin remodelling. Reporting in Cell, Palamidessi et al. show that Rab5-dependent endocytosis is a requirement for spatially-restricted Rac activation that occurs downstream of RTKs. The delivery of active Rac to specific plasma membrane regions ensures the formation of the actin protrusions that are important for directed migration.

Rab5 is a known regulator of endocytosis. Ectopic expression of Rab5 in HeLa cells led to increased levels of active Rac as well as inducing the formation of both lamellipodia and circular dorsal ruffles (Fig. 1A). Following treatment of cells with hepatocyte growth factor (HGF) the authors observed that both ruffle formation and Rac activation were dependent on Rab5, suggesting that Rac activation by RTKs also requires Rab5.

Ruffle formation and Rac activation were impaired when clathrin-mediated endocytosis was blocked. Furthermore, activated Rac localised at early endosomes upon Rab 5 transfection, showing that the endosomal membrane is the primary site for Rac activation by Rab5. But how is Rac activated on endosomes? The authors found that silencing the Rac guanine exchange factor (GEF) Tiam1 abrogated Rac activation and that localisation of Tiam1 at endosomal membranes was Rab5-dependent too. Thus, Rab5 appears to regulate the trafficking of Rac and Tiam1 to endosomes, which leads to the activation of Rac at that location.

Active Rac was recycled back to the plasma membrane at sites of membrane ruffling. Blocking the ability of Rac to recycle impaired Rab5- and HGF-dependent actin dynamics, suggesting that endocytic trafficking is required to obtain spatial restriction of Rac signalling, leading to actin remodelling and the formation of specialized cellular protrusions.

Palamidessi et al. also provide evidence that Rab5 regulates both cell morphology and motility. When Rab5 was ectopically expressed in HeLa cells they changed to an elongated shape with multiple cell protrusions — this also resulted in the redistribution of integrin 3 and enhanced cell invasion in 3D matrices (Fig. 1B). But does the Rab5-to-Rac circuitry affect physiological or pathological processes? Individual tumour cells can adopt two different types of migration — mesenchymal or amoeboid. Switching between one type and the other confers some advantage during dissemination enhancing the ability of cells to plastically adapt to the tissue microenvironment. Rab5 expression promoted the conversion of melanoma cells to the mesenchymal migration, whereas Rab5 inhibition converted colon carcinoma cells to the amoeboid type, suggesting that Rab5 is required to regulate the tumour cell motility. Interestingly, Rab5 was also found to regulate primordial germ cell migration during zebrafish development. Thus, Rab5-to-Rac circuitry appears to be important for the regulation of cell motility both in vitro and in vivo.

Author: Kim Baumann