PDGF-regulated rab4-dependent recycling of alpha v beta 3 integrin from early endosomes is necessary for cell adhesion and spreading

Background: It has been postulated that the regulation of integrin vesicula r traffic facilitates cell migration by internalizing integrins at the rear of the cell and transporting them forward within vesicles for exocytosis a t the leading edge to form new contacts with the extracellular matrix. The rab family of GTPases control key targeting events in the endo/exocytic pat hway; therefore, these GTPases may be involved in the regulation of cell-ma trix contact assembly. Results: The endo/exocytic cycle of (alphav beta3 and alpha5 beta1 integrin s was studied using mouse 3T3 fibroblast cell lines. In serum-starved cells , internalized integrins were transported through rab4-positive, early endo somes and arrived at the rab11-positive, perinuclear recycling compartment approximately 30 min after endocytosis. From the recycling compartment, int egrins were recycled to the plasma membrane in a rab11-dependent fashion. F ollowing treatment with PDGF, (alphav beta3 integrin, but not alpha5 beta1, was rapidly recycled directly back to the plasma membrane from the early e ndosomes via a rab4-dependent mechanism without the involvement of rab11. T his rapid recycling pathway directed alphav beta3 to numerous small puncta distributed evenly across the dorsal surface of the cell, and the integrin only became localized into focal complexes at later times following PDGF ad dition. Interestingly, inhibition of PDGF-stimulated alphav beta3 recycling using dominant-negative rab4 mutants compromised cell adhesion and spreadi ng on vitronectin (a ligand for (alphav beta3), but adhesion to fibronectin (a ligand for (alpha5 beta1 and (alphav beta3) was unchanged. Conclusions: We propose that growth factor-regulated, rab4-dependent recycl ing of (alphav beta3 integrin from early endosomes to the plasma membrane i s a critical upstream event in the assembly of cell-matrix contacts.