Journal
BIOCHEMICAL JOURNAL
Volume 475, Issue -, Pages 1-22Publisher
PORTLAND PRESS LTD
DOI: 10.1042/BCJ20170802
Keywords
-
Categories
Funding
- Michael J. Fox Foundation for Parkinson's research [12938]
- Medical Research Council [MC_UU_12016/2, MR/P00704X/1]
- AstraZeneca
- Boehringer-Ingelheim
- GlaxoSmithKline
- Merck KGaA
- Janssen Pharmaceutica
- Pfizer
- Henry G. Leong Endowed Professorship in Neurology
- Medical Research Council [MC_UU_00018/1, MC_U127070193, MR/P00704X/1, MC_UU_12016/2, G0700656] Funding Source: researchfish
- Parkinson's UK [H-1701] Funding Source: researchfish
- MRC [MC_UU_12016/2, MC_U127070193, MR/P00704X/1, MC_UU_00018/1, G0700656] Funding Source: UKRI
Ask authors/readers for more resources
Mutations that activate the LRRK2 (leucine-rich repeat protein kinase 2) 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 the present study, we analyse the selectivity and sensitivity of polyclonal and monoclonal phospho-specific antibodies raised against nine 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 the present study will help with the assessment of LRRK2 activity and examination of which Rab proteins are phosphorylated in vivo. These antibodies could also be used to assess the impact of LRRK2 inhibitors in future clinical trials.
Authors
I am an author on this paper
Click your name to claim this paper and add it to your profile.
Reviews
Recommended
No Data Available