A yeast-two hybrid screen with Lyn identified a novel 130 kDa multidomain protein with a 36% identity to Actin Filament Associated Protein (AFAP) 110 and similar domains, including PH domains, potential sites of tyrosine and serine/threonine phosphorylation, a leucine-zipper domain, a potential actin binding site and multimerization site. AFAP110 has been shown to have a role in modulating actin filament integrity and induce lamellipodia formation, and is known to interact with Src family kinases. The aim of this thesis was to characterize this novel protein named Lyn-Associated Cytoskeletal Modulator (LACM) and determine any molecular interactions in order to attempt to elucidate a role for the protein in cell signaling through Lyn. LACM is encoded by a gene consisting of 18 exons and is located on human chromosome 5q33.1 and mouse chromosome 18 E1. LACM protein is expressed through a number of cell types including the R11 erythroid cell line, and mouse tissues including brain, lung, heart and embryos. LACM was shown to multimerize, and subcellular localization of the protein was observed to concentrate around the cell membrane at sites of filamentous actin in filopodia, lamellipodia and stress fibres. The carboxy-terminus of LACM was observed to localize the protein to sites at the cell membrane and through the cytoplasm. Removal of this terminal region resulted in all LACM protein localizing to the nucleus in punctuate spots. LACM protein was observed in heart muscle and potentially has a role at sites of nerve junctions on cardiac myocytes. LACM was shown to interact with the SH3 domain of Lyn at a polyproline motif on LACM. LACM was observed to co-localize and co-immunoprecipitate with Lyn and was tyrosine phosphorylated by the kinase domain of Lyn. Interestingly, the consititutively active Lyn and LACM caused transfected cells to
| Identifer | oai:union.ndltd.org:ADTP/258967 |
| Date | January 2009 |
| Creators | McCarthy, David James |
| Publisher | University of Western Australia. School of Medicine and Pharmacology, Western Australian Institute for Medical Research |
| Source Sets | Australiasian Digital Theses Program |
| Language | English |
| Detected Language | English |
| Rights | Copyright David McCarthy, http://www.itpo.uwa.edu.au/UWA-Computer-And-Software-Use-Regulations.html |
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