Use this URL to cite or link to this record in EThOS: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.654616
Title: A direct interaction between the Parkinson's disease protein leucine-rich repeat kinase 2 and specific β-tubulin isoforms regulates tubulin acetylation
Author: Spain, V.
ISNI:       0000 0004 5359 0905
Awarding Body: University College London (University of London)
Current Institution: University College London (University of London)
Date of Award: 2015
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Abstract:
Mutations in LRRK2 are a common cause of Parkinson’s disease (PD). LRRK2 encodes leucine-rich repeat kinase 2 (LRRK2), a ROCO protein. It has an enzymatic core consisting of a Ras of complex proteins (Roc) GTPase domain and kinase domain, surrounded by protein-protein interaction regions. Pathogenic LRRK2 mutations modify activity in these enzymatic domains, but how this leads to neurodegeneration is still to be elucidated. One of the few confirmed LRRK2 interactors is tubulin, the main constituent of microtubules (MTs) and part of the cytoskeletal network. Disease-causing mutations in LRRK2 alter this network, reducing neurite outgrowth and leading to accumulation of hyperphosphorylated MT-associated protein (MAP) tau. Meanwhile changes in post-translational modifications of tubulin and MAPs alter the dynamic instability of MTs, leading to aberrant axonal transport, synaptic dysfunction and axonal degeneration. I investigated the LRRK2-tubulin interaction. Using yeast two-hybrid I demonstrated that the interaction is conferred by the LRRK2 Roc domain and the C-terminus of the β-tubulin isoforms TUBB, TUBB4 and TUBB6. The interaction requires Lys362 and Ala364 and is blocked in isoforms expressing a serine at these positions. This site is on the luminal face of MT protofibrils, close to the paclitaxel binding site and α-tubulin Lys40 acetylation site, both of which are involved in MT stability. This location is poorly accessible within mature, stabilised MTs but exposed in dynamic MT populations. Consistent with this finding, endogenous LRRK2 located to dynamic growth cone MTs in SH-SY5Y cells. Overexpression and knock-out studies in HEK cells and mouse embryonic fibroblasts showed that LRRK2 is associated with reduced α-tubulin acetylation. These results demonstrate the specificity of the LRRK2-tubulin interaction, suggesting LRRK2 distribution at the cytoskeleton is determined by the tubulin composition and may vary between cell types. Changes in MT acetylation in the presence of disease-causing LRRK2 mutations could contribute to pathogenic mechanisms, with altered MT stability implicated in PD neurodegeneration. As mutations affecting the β-tubulin C-terminal residues could disrupt the LRRK2 interaction without compromising MT integration, a cohort of late-onset familial PD cases was also screened for mutations within the cytoskeleton.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.654616  DOI: Not available
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