Lysosomal positioning regulates Rab10 phosphorylation at LRRK2-positive lysosomes

Abstract

Genetic variation at the Leucine-rich repeat kinase 2 (LRRK2) locus contributes to risk of familial and sporadic Parkinson disease. Recent data have shown a robust association between localization to various membranes of the endolysosomal system and LRRK2 activation. However, the mechanism(s) underlying LRRK2 activation at endolysosomal membranes are still poorly understood. Here we artificially direct LRRK2 to six different membranes within the endolysosomal system. We demonstrate that LRRK2 is activated and able to phosphorylate three of its Rab substrates (Rab10, Rab12 and Rab29) at each compartment. However, we report differing localization of pRab10 and pRab12 at the lysosomal and Golgi membranes. Specifically, we found that pRab10 colocalizes with a sub-population of perinuclear LRRK2-positive Golgi/lysosomal compartments whereas pRab12 localized to all LRRK2-positive Golgi/lysosomal membranes across the cell. When organelle positioning is manipulated by sequestering lysosomes to the perinuclear area, pRab10 colocalization with LRRK2 significantly increases. We also show recruitment of JIP4, a pRab10 effector that we have recently linked to LYTL, after trapping LRRK2 at various membranes. Taken together, we demonstrate that the association of LRRK2 to membranous compartments is sufficient for its activation and Rab phosphorylation independent of membrane identity. Our system also identifies a potential mechanism underlying the distinct relationships between LRRK2 and its substrates Rab10 and Rab12.

Development of a multiplexed targeted mass spectrometry assay for LRRK2-phosphorylated Rabs and Ser910/Ser935 biomarker sites

“…Is it also possible that differences in effector binding affinity could influence the steady state phospho-Rab1 levels in cells by competing with kinases and phosphatases targeting the Switch-II site. The latter point is also likely to be relevant for other Rab proteins and warrants further analysis [ 46 ].…”

Preclinical modeling of chronic inhibition of the Parkinson’s disease associated kinase LRRK2 reveals altered function of the endolysosomal system in vivo

“…Lastly, we recently reported differing localization patterns between pS106 Rab12 and pT73 Rab10 when artificially trapping LRRK2 to different cellular membranes. Independent of which membrane LRRK2 was directed to intracellularly, pRab12 was found to colocalize with LRRK2, whereas pRab10 was primarily detected at perinuclear lysosomes and Golgi after directing LRRK2 to these membranes [ 38 ]. This falls in line with the idea that a more complex mechanism surrounds the Rab10:LRRK2 interaction and thus any influence LRRK2 has on Rab10 is more conditional compared to Rab12.…”

Increased LRRK2 kinase activity alters neuronal autophagy by disrupting the axonal transport of autophagosomes

“…14,16 Recent studies found that membrane association, but not membrane identity, is important for LRRK2 activation by Rab29. 15,51 Since we detected Rab29 on axonal AVs, we investigated whether overexpression of Rab29 affects the processivity of AV transport in a similar manner to the LRRK2-G2019S mutation. We transiently expressed EGFP-Rab29 and mCherry-LC3 in WT or G2019S KI mouse cortical neurons.…”

“…JIP3 and JIP4 are motor adaptor proteins that bind LRRK2phosphorylated Rab proteins: Rab8, Rab10, and Rab35. 11,12,51 In astrocytes, expression of LRRK2-G2019S increased recruitment of JIP4 to damaged lysosomes by enhancing Rab phosphorylation. 12 In line with these findings, we detected significantly higher levels of JIP4 on autophagosomes isolated from the brains of G2019S KI mice than WT mice (Figure 6A).…”