Membrane targeting activates Leucine-rich repeat kinase 2 with differential effects on downstream Rab activation

Abstract

Genetic variation at the Leucine-rich repeat kinase 2 (LRRK2) locus contributes to enhanced lifetime risk of familial and sporadic Parkinsons disease. Previous data have demonstrated that recruitment to various membranes of the endolysosomal system results in LRRK2 activation. However, the mechanism(s) underlying LRRK2 activation at endolysosomal membranes and the cellular consequences of these events are still poorly understood. Here, we directed LRRK2 to lysosomes and early endosomes, triggering both LRRK2 autophosphorylation and phosphorylation of the direct LRRK2 substrates Rab10 and Rab12. However, when directed to the lysosomal membrane, pRab10 was restricted to perinuclear lysosomes, whereas pRab12 was visualized on both peripheral and perinuclear LRRK2-positive lysosomes, suggesting that lysosomal positioning provides additional regulation of LRRK2-dependent Rab phosphorylation. Anterograde transport of lysosomes to the cell periphery by increasing expression of ARL8B and SKIP or by knockdown of the motor adaptor protein JIP4 blocked recruitment and phosphorylation of Rab10 by LRRK2. Conversely, overexpression of the Rab7 effector protein RILP resulted in lysosomal clustering within the perinuclear area and increased LRRK2-dependent Rab10 recruitment and phosphorylation. The regulation of Rab10 phosphorylation in the perinuclear area depends on counteracting phosphatases, as knockdown of phosphatase PPM1H significantly increased pRab10 signal and lysosomal tubulation in the perinuclear region. Our novel findings suggest LRRK2 can be activated at multiple cellular membranes including lysosomes, and that lysosomal positioning further provides regulation of some Rab substrates likely via differential phosphatase activity in nearby cellular compartments.