Cell migration is an essential physiological process required for embryogenesis, tissue repair and immune surveillance. Directional cell migration requires coordinated regulation of multiple integrin-mediated cellular processes, including dynamic regulation of the actin cytoskeleton, precise control of membrane protrusion events and the constant recycling of adhesion receptors. While it is clear that regulation of the small guanosine triphosphatases (GTPases) Rac1, Arf6 and RhoA is critical for these processes, the integrin-dependent mechanisms responsible for cyclic activation and dynamic coordination of GTPase signalling are only partially understood. Here, analysis of three published mass spectrometry (MS) studies cataloguing integrin-dependent adhesion complexes identified filamin-A and IQGAP1 as potential candidates linking β1 integrin to the regulation of Rac1 activity. Using immunoprecipitation, MS analysis, immunocytochemistry and RNAi, filamin-A and IQGAP1 were found to be recruited to integrin activation sites, where they constrained Rac1 activity via the recruitment of the GTPase-activating protein RacGAP1. The functional relevance of Rac1 deactivation, through a RacGAP1 and IQGAP1-mediated mechanism, is to permit efficient membrane protrusion and directional cell migration. Subsequently, IQGAP1 was identified as a molecule co-ordinating Rac1 and Arf6 activities downstream of β1 integrin engagement, via the recruitment of the GTPase activity modulators RacGAP1, srGAP2 and HERC1. This lead us to propose a model whereby IQGAP1, through the recruitment of multiple small GTPase activity modulators, co-ordinates the two small GTPases Rac1 and Arf6, to efficiently regulate directional cell migration. Dyregulated cell migration due to integrin over-activation is associated with tumour invasion. Increased recycling of α5β1 integrin, resulting from expression of mutant p53 or inhibition of αVβ3 integrin function, leads to random cell migration on 2D substrates and promotes tumour invasion via activation of the pro-invasive kinase Akt. Here, the RacGAP1- IQGAP1 complex was identified as a key component of this pathway. In particular, RacGAP1 was found to be phosphorylated by Akt2 on T249, a phosphorylation event that promoted RacGAP1 recruitment to IQGAP1, at the cell front, and triggered cell invasion by inducing a Rac1/RhoA activity switch. These findings demonstrated that Akt activation, downstream of α5β1 integrin recycling, promotes fibronectin-mediated cell invasion by activating a novel RacGAP1/IQGAP1/Rac1/RhoA pathway. Taken together, we identified a novel signalling nexus, downstream of integrin activation and/or recycling that co-ordinate the small GTPases Rac1, Arf6 and RhoA during cell migration and invasion.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:574366 |
| Date | January 2013 |
| Creators | Jacquemet, Guillaume |
| Contributors | Humphries, Martin; Tournier, Cathy |
| Publisher | University of Manchester |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | https://www.research.manchester.ac.uk/portal/en/theses/integrinmediated-regulation-of-small-gtpases(dac891db-1c6d-4769-ba15-b24fef4e46b3).html |
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