The role of WASP family members in Dictyostelium discoideum cell migration

Davidson, Andrew J. (2014) The role of WASP family members in Dictyostelium discoideum cell migration. PhD thesis, University of Glasgow.

Abstract

The WASP family of proteins are nucleation-promoting factors that dictate the temporal and spatial dynamics of Arp2/3 complex recruitment, and hence actin polymerisation. Consequently, members of the WASP family, such as SCAR/WAVE and WASP, drive processes such as pseudopod formation and clathrin-mediated endocytosis, respectively. However, the nature of functional specificity or overlap of WASP family members is controversial and also appears to be contextual. For example, some WASP family members appear capable of assuming each other’s roles in cells that are mutant for certain family members. How the activity of each WASP family member is normally limited to promoting the formation of a specific subset of actin-based structures and how they are able to escape these constraints in order to substitute for one another, remain unanswered questions. Furthermore, how the WASP family members collectively contribute to complex processes such as cell migration is yet to be addressed.
To examine these concepts in an experimentally and genetically tractable system we have used the single celled amoeba Dictyostelium discoideum. The regulation of SCAR via its regulatory complex was investigated by dissecting the Abi subunit. Abi was found to be essential for complex stability but not for its recruitment to the cell cortex or its role in pseudopod formation. The roles of WASP A were examined by generating a wasA null strain. Our results contradicted previous findings suggesting that WASP A was essential for pseudopod formation and instead demonstrated that WASP A was required for clathrin-mediated endocytosis. Unexpectedly, WASP A – driven clathrin-mediated endocytosis was found to be necessary for efficient uropod retraction during cell migration and furrowing during cytokinesis. Finally, we created a double scrA/wasA mutant, and found that it was unable to generate pseudopodia. Therefore, we were able to confirm that SCAR is the predominant driver of pseudopod formation in wild-type Dictyostelium cells, and that only WASP A can assume its role in the scrA null. Surprisingly, the double mutant was also deficient in bleb formation, showing that these proteins are also necessary for this alternative, Arp2/3 complex-independent mode of motility. This implies that there exists interplay between the different types of actin-based protrusions and the molecular pathways that underlie their formation.

• Item Type: Thesis (PhD)
• Qualification Level: Doctoral
• Keywords: WASP, SCAR, WAVE, Abi, Arp2/3, actin, clathrin, myosin, pseudopod, bleb, filopod, protrusion, migration, Dictyostelium, motility, chemotaxis, cytokinesis
• Subjects: Q Science > QH Natural history > QH301 Biology; Q Science > QH Natural history > QH426 Genetics
• Colleges/Schools: College of Medical Veterinary and Life Sciences > School of Cancer Sciences > Beatson Institute of Cancer Research
• Supervisor's Name: Insall, Professor Robert H.
• Date of Award: 2014
• Depositing User: Mr Andrew Davidson
• Unique ID: glathesis:2014-4963
• Copyright: Copyright of this thesis is held by the author.
• Date Deposited: 15 Apr 2014 10:30
• Last Modified: 15 Apr 2014 10:55
• URI: https://theses.gla.ac.uk/id/eprint/4963