The Parkinson's disease VPS35[D620N] mutation enhances LRRK2-mediated Rab protein phosphorylation in mouse and human.

Rafeeq Mir,Christoph Hotzy,Thomas Brücke,Nicole K Polinski,Pawel Lis,Theresa König,Alexander Zimprich,Meng-Yun Chou,Esther Sammler,Dario R Alessi,Thomas Macartney,Ivan Milenkovic,Andrew J.M Howden,Francesca Tonelli,Terina N Martinez

doi:10.1042/bcj20180248

Abstract

Missense mutations in the LRRK2 (Leucine-rich repeat protein kinase-2) and VPS35 genes result in autosomal dominant Parkinson's disease. The VPS35 gene encodes for the cargo-binding component of the retromer complex, while LRRK2 modulates vesicular trafficking by phosphorylating a subgroup of Rab proteins. Pathogenic mutations in LRRK2 increase its kinase activity. It is not known how the only thus far described pathogenic VPS35 mutation, [p.D620N] exerts its effects. We reveal that the VPS35[D620N] knock-in mutation strikingly elevates LRRK2-mediated phosphorylation of Rab8A, Rab10, and Rab12 in mouse embryonic fibroblasts. The VPS35[D620N] mutation also increases Rab10 phosphorylation in mouse tissues (the lung, kidney, spleen, and brain). Furthermore, LRRK2-mediated Rab10 phosphorylation is increased in neutrophils as well as monocytes isolated from three Parkinson's patients with a heterozygous VPS35[D620N] mutation compared with healthy donors and idiopathic Parkinson's patients. LRRK2-mediated Rab10 phosphorylation is significantly suppressed by knock-out or knock-down of VPS35 in wild-type, LRRK2[R1441C], or VPS35[D620N] cells. Finally, VPS35[D620N] mutation promotes Rab10 phosphorylation more potently than LRRK2 pathogenic mutations. Available data suggest that Parkinson's patients with VPS35[D620N] develop the disease at a younger age than those with LRRK2 mutations. Our observations indicate that VPS35 controls LRRK2 activity and that the VPS35[D620N] mutation results in a gain of function, potentially causing PD through hyperactivation of the LRRK2 kinase. Our findings suggest that it may be possible to elaborate compounds that target the retromer complex to suppress LRRK2 activity. Moreover, patients with VPS35[D620N] associated Parkinson's might benefit from LRRK2 inhibitor treatment that have entered clinical trials in humans.

Highlights

Parkinson’s disease (PD) is a common neurodegenerative condition with numbers expected to rise [1]
This is based on the following findings: the pathogenic, PD causing VPS35[D620N] knock-in mutation stimulates Leucine-Rich Repeat protein kinase-2 (LRRK2)-mediated phosphorylation of at least 3 Rab proteins (Rab8A, Rab10, and Rab12) in mouse embryonic fibroblasts (MEFs) (Figure 2) as well as enhancing Rab10 phosphorylation in mouse tissues that express LRRK2 (Figure 4)
In MEFs, we demonstrate that the VPS35[D620N] knock-in mutation markedly enhances autophosphorylation of LRRK2 at Ser1292, consistent with LRRK2 becoming activated (Figure 3B)

Summary

Introduction

Parkinson’s disease (PD) is a common neurodegenerative condition with numbers expected to rise [1]. Most PD cases are idiopathic, studying the disease mechanisms of the rarer genetic PD forms is expected to advance our knowledge in the aetiology of idiopathic disease and help identify targets for the development of biomarker and novel treatments. That mutations in LRRK2 (Leucine-rich repeat protein kinase-2) result in PD was first described in 2004 [4,5] and are one of the most common genetic causes for PD. The G2019S mutation directly increases the kinase activity of recombinant LRRK2 [9], while the R1441G/C and Y1699C mutations do so indirectly by promoting the recruitment of LRRK2 to the Rab protein located at the Golgi, leading to LRRK2 activation through an as yet unknown mechanism [10,11,12]

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Results

Conclusion