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Title: Regulation of N-WASP-dependent actin polymerisation
Author: Humphries, A. C.
Awarding Body: University College London (University of London)
Current Institution: University College London (University of London)
Date of Award: 2013
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N-WASP activates the Arp2/3 complex to stimulate actin polymerisation during a number of cellular processes. These include, phagocytosis, endocytosis, and invadopodia and podosome formation. In addition, it is also frequently recruited by pathogens to assist in their actin-dependent spread. During its egress from the cell, vaccinia virus fuses with the plasma membrane and promotes the activation of Src and Abl family kinases. This event leads to phosphorylation of tyrosine 112 and tyrosine 132 of the viral transmembrane protein, A36, triggering a signalling cascade that stimulates actin polymerisation. The phospho-tyrosine residues provide docking sites for the cellular adaptors Nck and Grb2, with Nck recruiting WIP-N-WASP and in turn the Arp2/3 complex. I found that clathrin is transiently recruited to vaccinia in the moments before actin polymerisation. Clathrin performs an organisational role, clustering A36 beneath the virus to enhance robust actin tail formation. Loss of clathrin recruitment resulted in a more disperse localisation of A36 and N-WASP, which led to an altered actin tail morphology, indicating that the spatial organisation of N-WASP is a parameter affecting actin polymerisation. During actin tail formation, I additionally found that Cdc42 could regulate N-WASP activity. Cdc42 supported the main signalling nexus of Nck-WIP-N-WASP, with its major role attributed to stabilising N-WASP beneath the viral particle. I further found that Cdc42 acts in a feed-forward loop with N-WASP and the RhoGEF intersectin-1. This pathway is conserved in FcγR-mediated phagocytosis, validating the use of pathogens to understand the molecular detail of Arp2/3-dependent actin-signalling pathways.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available