Title: The role of IQGAP1 in actin polymerisation : possible implications in motility
Abstract: IQGAP1 is a large (189 kDa) modular protein, discovered in 1994 as a multipartner protein associating with the signalling proteins calmodulin (CaM), Cdc42, Rac1, and directly with actin. It is involved in the organisation of actin and microtubules (MT) in motility processes such as adhesion, migration and cytokinesis, and is overexpressed especially in metastatic cancers. Ln cells IQGAP1 is found mainIy in the lamellipodium, colocalising with N-WASP, and in adherens junctions. IQGAP1 also binds proteins associated with MT plus ends, and it is involved in several signalling pathways, among others the MEK-ERK signalling. The molecular mechanisms responsible for the activities of IQGAP1 are not known, in part due to the large size, poor solubility and unknown folding of the protein, the large number of its ligands and the lack of appropriate methods to challenge putative activities. Recent work from the laboratoiries of G. Bloom and R. Kroschewski, with our collaboration, has shown that IQGAP1 stimulates actin assembly through the N-WASP-Arp2/3 pathway. I have expressed and purified the full length protein and several fragments. I show that full length IQGAP1 is a potent barbed end capping protein, that blocks assembly of actin at the filament barbed end with a Kd of 34 nM and disassembly of ADP-F-actin with a Kd of 4 nM. The barbed end capping activity is abolished by CaM (Kd CaM-IQGAP1 = 40 nM). I show that CaM abolishes the interaction of IQGAP1 with not onIy Cdc42 but also Rac1 and the BR-CRIB domain of N-WASP. I further challenge IQGAP1's N-WASP activating abilities. These results provide new insight into the complex activities of IQGAP1 and their regulation by CaM.
Publication Year: 2010
Publication Date: 2010-01-01
Language: en
Type: dissertation
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