Integrin-mediated regulation of small GTPases

Abstract

Cell migration is an essential physiological process required for embryogenesis,tissue repair and immune surveillance. Directional cell migration requires coordinatedregulation of multiple integrin-mediated cellular processes, including dynamic regulation ofthe actin cytoskeleton, precise control of membrane protrusion events and the constantrecycling of adhesion receptors. While it is clear that regulation of the small guanosinetriphosphatases (GTPases) Rac1, Arf6 and RhoA is critical for these processes, theintegrin-dependent mechanisms responsible for cyclic activation and dynamiccoordination of GTPase signalling are only partially understood.Here, analysis of three published mass spectrometry (MS) studies cataloguingintegrin-dependent adhesion complexes identified filamin-A and IQGAP1 as potentialcandidates linking β1 integrin to the regulation of Rac1 activity. Usingimmunoprecipitation, MS analysis, immunocytochemistry and RNAi, filamin-A andIQGAP1 were found to be recruited to integrin activation sites, where they constrainedRac1 activity via the recruitment of the GTPase-activating protein RacGAP1. Thefunctional relevance of Rac1 deactivation, through a RacGAP1 and IQGAP1-mediatedmechanism, is to permit efficient membrane protrusion and directional cell migration.Subsequently, IQGAP1 was identified as a molecule co-ordinating Rac1 and Arf6 activitiesdownstream of β1 integrin engagement, via the recruitment of the GTPase activitymodulators RacGAP1, srGAP2 and HERC1. This lead us to propose a model wherebyIQGAP1, through the recruitment of multiple small GTPase activity modulators,co-ordinates the two small GTPases Rac1 and Arf6, to efficiently regulate directional cellmigration.Dyregulated cell migration due to integrin over-activation is associated with tumourinvasion. Increased recycling of alpha5β1 integrin, resulting from expression of mutant p53 orinhibition of alphaVβ3 integrin function, leads to random cell migration on 2D substrates andpromotes tumour invasion via activation of the pro-invasive kinase Akt. Here, theRacGAP1- IQGAP1 complex was identified as a key component of this pathway. Inparticular, RacGAP1 was found to be phosphorylated by Akt2 on T249, a phosphorylationevent that promoted RacGAP1 recruitment to IQGAP1, at the cell front, and triggered cellinvasion by inducing a Rac1/RhoA activity switch. These findings demonstrated that Aktactivation, downstream of alpha5β1 integrin recycling, promotes fibronectin-mediated cellinvasion by activating a novel RacGAP1/IQGAP1/Rac1/RhoA pathway.Taken together, we identified a novel signalling nexus, downstream of integrin activationand/or recycling that co-ordinate the small GTPases Rac1, Arf6 and RhoA during cellmigration and invasion.

Date of Award	1 Aug 2013
Original language	English
Awarding Institution	The University of Manchester
Supervisor	Martin Humphries (Supervisor) & Cathy Tournier (Supervisor)