Parkinson disease-associated mutations in LRRK2 cause centrosomal defects via Rab8a phosphorylation

Abstract

BackgroundMutations in LRRK2 are a common genetic cause of Parkinson’s disease (PD). LRRK2 interacts with and phosphorylates a subset of Rab proteins including Rab8a, a protein which has been implicated in various centrosome-related events. However, the cellular consequences of such phosphorylation remain elusive.MethodsHuman neuroblastoma SH-SY5Y cells stably expressing wildtype or pathogenic LRRK2 were used to test for polarity defects in the context of centrosomal positioning. Centrosomal cohesion deficits were analyzed from transiently transfected HEK293T cells, as well as from two distinct peripheral cell types derived from LRRK2-PD patients. Kinase assays, coimmunoprecipitation and GTP binding/retention assays were used to address Rab8a phosphorylation by LRRK2 and its effects in vitro. Transient transfections and siRNA experiments were performed to probe for the implication of Rab8a and its phosphorylated form in the centrosomal deficits caused by pathogenic LRRK2.ResultsHere, we show that pathogenic LRRK2 causes deficits in centrosomal positioning with effects on neurite outgrowth, cell polarization and directed migration. Pathogenic LRRK2 also causes deficits in centrosome cohesion which can be detected in peripheral cells derived from LRRK2-PD patients as compared to healthy controls, and which are reversed upon LRRK2 kinase inhibition. The centrosomal cohesion and polarity deficits can be mimicked when co-expressing wildtype LRRK2 with wildtype but not phospho-deficient Rab8a. The centrosomal defects induced by pathogenic LRRK2 are associated with a kinase activity-dependent increase in the centrosomal localization of phosphorylated Rab8a, and are prominently reduced upon RNAi of Rab8a.ConclusionsOur findings reveal a new function of LRRK2 mediated by Rab8a phosphorylation and related to various centrosomal defects.Electronic supplementary materialThe online version of this article (10.1186/s13024-018-0235-y) contains supplementary material, which is available to authorized users.

The Emerging Functions of LRRK2 and Rab GTPases in the Endolysosomal System

“…The subsequent and systematic analyses demonstrated that Rab3a-d, Rab5a-c, Rab8a/b, Rab10, Rab12, Rab29 (also known as Rab7L1), Rab35 and Rab43 are phosphorylated by LRRK2 at least upon overexpression (Steger et al, 2017). Other groups have also reported that Rab8, Rab10 and Rab29 behave as excellent substrates of LRRK2 in cells (Fujimoto et al, 2018;Liu et al, 2018;Madero-Perez et al, 2018a). At endogenous levels, LRRK2-mediated phosphorylation likely occurs on Rab3a-d, Rab8a/b, Rab10, Rab12, Rab35 and Rab43 (Steger et al, 2017).…”

Vesicular Dysfunction and the Pathogenesis of Parkinson’s Disease: Clues From Genetic Studies

“…Similarly, Rab8A has also been shown to play a role in membrane trafficking and clearance as well as protein transport (Chung et al, 2017). Further evidence for the role of Rab8A in development and pathogenesis comes as studies have shown that LRRK2 phosphorylation of Rab8A can cause centrosomal defects and thus cause widespread effects on neurite growth and migration (Madero-Pérez et al, 2018a).…”

LRRK2 is recruited to phagosomes and co-recruits Rab8 and Rab10 in human pluripotent stem cell-derived macrophages

“…vesicle transport and membrane recycling to the cell surface(Banton et al, 2014), autophagy (with the ALS-related gene, C9orf72, implicated as a Rab8 GEF) (Corbier and Sellier, 2017), and has previously been linked to LRRK2 in, variously, endolysosomal trafficking(Rivero-Ríos et al, 2019), centrosome function(Madero-Pérez et al, 2018) and…”