Lis, P;
Burel, S;
Steger, M;
Mann, M;
Brown, F;
Diez, F;
Tonelli, F;
... Alessi, DR; + view all
(2018)
Development of phospho-specific Rab protein antibodies to monitor in vivo activity of the LRRK2 Parkinson's disease kinase.
Biochemical Journal
, 475
(1)
pp. 1-22.
10.1042/BCJ20170802.
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Abstract
Mutations that activate the LRRK2 protein kinase, predispose to Parkinson's disease, suggesting that LRRK2 inhibitors might have therapeutic benefit. Recent work has revealed that LRRK2 phosphorylates a subgroup of 14 Rab proteins, including Rab10, at a specific residue located at the centre of its effector binding Switch-II motif. In this study, we analyse the selectivity and sensitivity of polyclonal and monoclonal phospho-specific antibodies raised against 9 different LRRK2 phosphorylated Rab proteins (Rab3A/3B/3C/3D, Rab5A/5B/5C, Rab8A/8B, Rab10, Rab12, Rab29[T71], Rab29[S72], Rab35 and Rab43). We identify rabbit monoclonal phospho-specific antibodies (MJFF-pRAB10) that are exquisitely selective for LRRK2 phosphorylated Rab10, detecting endogenous phosphorylated Rab10 in all analysed cell lines and tissues, including human brain cingulate cortex. We demonstrate that the MJFF-pRAB10 antibodies can be deployed to assess enhanced Rab10 phosphorylation resulting from pathogenic (R1441C/G or G2019S) LRRK2 knock-in mutations as well as the impact of LRRK2 inhibitor treatment. We also identify rabbit monoclonal antibodies displaying broad specificity (MJFF-pRAB8) that can be utilised to assess LRRK2 controlled phosphorylation of a range of endogenous Rab proteins including Rab8A, Rab10 and Rab35. The antibodies described in this study will help with assessment of LRRK2 activity and examination of which Rab proteins are phosphorylated in vivo. These antibodies could also be used to assess impact of LRRK2 inhibitors in future clinical trials.
| Type: | Article |
|---|---|
| Title: | Development of phospho-specific Rab protein antibodies to monitor in vivo activity of the LRRK2 Parkinson's disease kinase |
| Location: | England |
| Open access status: | An open access version is available from UCL Discovery |
| DOI: | 10.1042/BCJ20170802 |
| Publisher version: | http://doi.org/10.1042/BCJ20170802 |
| Language: | English |
| Additional information: | Copyright © 2018 The Author(s) https://creativecommons.org/licenses/by/4.0/ This is an open access article published by Portland Press Limited on behalf of the Biochemical Society and distributed under the Creative Commons Attribution License 4.0 (CC BY). |
| Keywords: | Parkinsons disease, Rab GTPases, Rab10, antibodies, leucine rich repeat kinase, protein kinase |
| UCL classification: | UCL UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Brain Sciences UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Brain Sciences > UCL Queen Square Institute of Neurology |
| URI: | https://discovery.ucl.ac.uk/id/eprint/10040587 |
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