Roles of lysosomotropic agents on LRRK2 activation and Rab10 phosphorylation

Abstract

Leucine-rich repeat kinase 2 (LRRK2), the major causative gene product of autosomal-dominant Parkinson's disease, is a protein kinase that phosphorylates a subset of Rab GTPases. Since pathogenic LRRK2 mutations increase its ability to phosphorylate Rab GTPases, elucidating the mechanisms of how Rab phosphorylation is regulated by LRRK2 is of great importance. We have previously reported that chloroquine-induced lysosomal stress facilitates LRRK2 phosphorylation of Rab10 to maintain lysosomal homeostasis. Here we reveal that Rab10 phosphorylation by LRRK2 is potently stimulated by treatment of cells with a set of lysosome stressors and clinically used lysosomotropic drugs. These agents commonly promoted the formation of LRRK2-coated enlarged lysosomes and extracellular release of lysosomal enzyme cathepsin B, the latter being dependent on LRRK2 kinase activity. In contrast to the increase in Rab10 phosphorylation, treatment with lysosomotropic drugs did not increase the enzymatic activity of LRRK2, as monitored by its autophosphorylation at Ser1292 residue, but rather enhanced the molecular proximity between LRRK2 and its substrate Rab GTPases on the cytosolic surface of lysosomes. Lysosomotropic drug-induced upregulation of Rab10 phosphorylation was likely a downstream event of Rab29 (Rab7L1)-mediated enzymatic activation of LRRK2. These results suggest a regulated process of Rab10 phosphorylation by LRRK2 that is associated with lysosomal overload stress, and provide insights into the novel strategies to halt the aberrant upregulation of LRRK2 kinase activity.

Parkinson’s VPS35[D620N] mutation induces LRRK2-mediated lysosomal association of RILPL1 and TMEM55B

“…Many previous studies have pointed toward lysosomal damage/dysfunction triggering LRRK2 pathway activation. For example, treatment of cells with lysosome-damaging agents such as chloroquine ( 58 , 59 ) or LLOMe ( 12 ) triggers LRRK2 recruitment to the lysosome, inducing its activation and enhanced phosphorylation of Rab8A and Rab10 on the lysosome. Chloroquine was also reported to induce relocalization of Rab8/Rab10 effectors EHBP1 and EHBP1L1 to the lysosome ( 58 ), whereas LLOMe induced lysosomal recruitment of the pRab10 motor adapter protein JIP4 ( 12 ) as well as the RAB7 GTPase-activating protein TBC1D15 ( 60 ).…”