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Rab10 Phosphorylation Detection by LRRK2 Activity Using SDS-PAGE with a Phosphate-binding Tag
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Rab10 Phosphorylation Detection by LRRK2 Activity Using SDS-PAGE with a Phosphate-binding Tag

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Roles of lysosomotropic agents on LRRK2 activation and Rab10 phosphorylation

Tomoki Kuwahara1, Kai Funakawa1, Tadayuki Komori1

  • 1Department of Neuropathology, Graduate School of Medicine, The University of Tokyo, Tokyo 113-0033, Japan.

Neurobiology of Disease
|September 12, 2020
PubMed

Insights

Lysosomal stress, induced by drugs, enhances Leucine-rich repeat kinase 2 (LRRK2) phosphorylation of Rab GTPases. This process, linked to Parkinson's disease, involves LRRK2 recruitment to lysosomes and Rab29 activation, offering new therapeutic insights.

Area of Science:

  • Neuroscience
  • Cell Biology
  • Biochemistry

Background:

  • Leucine-rich repeat kinase 2 (LRRK2) is a key protein kinase implicated in Parkinson's disease pathogenesis.
  • LRRK2 phosphorylates Rab GTPases, and mutations associated with Parkinson's disease enhance this activity.
  • Previous work showed chloroquine-induced lysosomal stress promotes LRRK2-Rab10 phosphorylation for lysosomal homeostasis.

Purpose of the Study:

  • To investigate how lysosomal stress and lysosomotropic drugs regulate LRRK2-mediated Rab GTPase phosphorylation.
  • To elucidate the mechanisms underlying the enhanced LRRK2 activity under lysosomal stress conditions.
  • To explore potential therapeutic strategies targeting LRRK2 dysregulation in Parkinson's disease.

Main Methods:

  • Cellular treatment with various lysosome stressors and clinically used lysosomotropic drugs.
  • Analysis of LRRK2 localization, Rab10 phosphorylation levels, and LRRK2 autophosphorylation (Ser1292).
  • Assessment of cathepsin B release and LRRK2-Rab GTPase proximity on lysosomes.
  • Investigation of the role of Rab29 (Rab7L1) in mediating LRRK2 activation.

Main Results:

  • Lysosomotropic drugs and lysosomal stressors potently stimulate LRRK2 phosphorylation of Rab10.
  • These agents induce LRRK2 recruitment to enlarged lysosomes and promote cathepsin B release.
  • LRRK2 kinase activity (Ser1292 autophosphorylation) was not increased, but LRRK2-Rab proximity on lysosomes was enhanced.
  • Rab10 phosphorylation upregulation is downstream of Rab29-mediated LRRK2 activation.

Conclusions:

  • LRRK2-mediated Rab10 phosphorylation is tightly regulated by lysosomal overload stress.
  • Lysosomotropic drugs enhance LRRK2-Rab GTPase interaction on lysosomes, independent of increased LRRK2 enzymatic activity.
  • Rab29 plays a crucial role in mediating LRRK2 activation under lysosomal stress.
  • Findings suggest novel strategies to modulate aberrant LRRK2 kinase activity in Parkinson's disease.
Keywords:
LRRK2LysosomeLysosomotropic agentsParkinson's diseasePhosphorylationRab

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