Use this URL to cite or link to this record in EThOS: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.652804
Title: Regulation of interactions in the dystrophin-associated protein complex by phosphorylation
Author: Ilsley, J. L.
Awarding Body: University of Edinburgh
Current Institution: University of Edinburgh
Date of Award: 2002
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Abstract:
It has recently been demonstrated that dystrophin, dystrobrevin and members of the sarcoglycan subcomplex are phosphorylated, and syntrophin and β-dystroglycan associate with signalling proteins such as nNOS and Grb2. The aim of this study was to identify novel phosphorylation events within the DAPC and determine their effects on the regulation of protein:protein interactions in the complex. Upon treatment of C2/C4 cells (a mouse muscle cell line) with peroxyvanadate, a potent tyrosine phosphatase inhibitor, β-dystroglycan is tyrosine-phosphorylated. This phosphorylation event was initially detected by mobility shifts on SDS-polyacrylamide gels, and confirmed by immunoprecipitation and two-dimensional gel electrophoresis. The potential functional significance of this tyrosine phosphorylation was investigated using peptide ‘SPOTs’ assays. Phosphorylation of tyrosine in the 15 most C-terminal residues of β-dystroglycan disrupts its interaction with dystrophin. The tyrosine residue in β-dystroglycan’s WW domain-binding motif PPPY appears to be the most crucial in disrupting the interaction. Therefore the β-dystroglycan/dystrophin interaction appears to be regulated by tyrosine phosphorylation. Various methods were used to investigate the protein:protein interactions of β-dystroglycan. In a yeast two-hybrid assay β-dystroglycan was found to interact with actin. This novel interaction was further characterised by F-actin sedimentation assays and it was demonstrated β-dystroglycan facilitated the sedimentation of F-actin during low speed centrifugation. Furthermore, electron microscopy studies revealed that β-dystroglycan bundles actin filaments. These studies highlight the importance of β-dystroglycan, not only in the regulation of the interaction between dystrophin and the rest of the DAPC but also its possible role in the regulation of the actin cytoskeleton. Understanding more about functions of dystrophin and the DAPC allows a greater insight into the pathogenesis of Duchenne muscular dystrophy.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.652804  DOI: Not available
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