Extracellular vesicle-mediated release of bis(monoacylglycerol)phosphate is regulated by LRRK2 and glucocerebrosidase activity.
Extracellular vesicle-mediated release of bis(monoacylglycerol)phosphate is regulated by LRRK2 and glucocerebrosidase activity.
Where did the research take place?
The study site has not been established. Author addresses may differ from where the research occurred.
Barcelona, ES · Author affiliation
Departament de Biomedicina, Unitat de Biologia Cel·lular, Facultat de Medicina i Ciències de la Salut, Centre de Recerca Biomèdica CELLEX, Institut d'Investigacions Biomèdiques August Pi i Sunyer, Universitat de Barcelona, Barcelona, Spain.Location evidence
Woburn, US · Author affiliation
NextCea Inc, Woburn, United States.Location evidence
US · Author affiliation · country only
Department of Biochemistry, Stanford University, Stanford, United States.Location evidence
Chicago, US · Author affiliation
Department of Neurology, Northwestern University, Chicago, United States.Location evidence
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Original abstract
The endolysosomal phospholipid bis(monoacylglycerol)phosphate (BMP) is aberrantly elevated in urine from Parkinson's patients carrying mutations in leucine-rich repeat kinase 2 (LRRK2) and glucocerebrosidase (GCase). Because BMP resides on, and regulates biogenesis of, endolysosomal intralumenal membranes that become extracellular vesicles (EVs) upon release, we hypothesized that increased urinary BMP reflects enhanced exocytosis of BMP-enriched EVs. We analyzed BMP metabolism and EV-associated BMP release in wild-type (WT) and R1441G LRRK2 mouse embryonic fibroblasts (MEFs). Immunofluorescence and transmission electron microscopy revealed structural alterations in endolysosomes and the antibody-accessible BMP pool, indicating disrupted endolysosomal homeostasis. Biochemical analysis of isolated EV fractions showed increased release of LAMP2-positive EVs by mutant cells, partially restored by LRRK2 kinase inhibition but further, variably, increased by GCase inhibition. Mass spectrometry detected higher total di-22:6-BMP and di-18:1-BMP in mutant LRRK2 MEFs compared to WT. Inhibition of LRRK2 partially restored cellular BMP, whereas GCase inhibition further elevated it. In EVs from mutant cells, LRRK2 inhibition reduced BMP content, while GCase inhibition tended to increase it. Metabolic labeling showed elevated BMP was not due to increased synthesis, despite higher levels of the BMP-synthesizing enzyme CLN5 in mutant MEFs and patient fibroblasts. Finally, pharmacological modulation of EV release and live total internal reflection fluorescence imaging in human G2019S LRRK2 fibroblasts further confirmed that BMP release is likely associated with EV secretion. Together, these results establish LRRK2 as a regulator of BMP in cells and its release through EVs and suggest that GCase activity further modulates this process in LRRK2 mutant cells. Mechanistic insights from these studies have implications for the use of BMP-positive EVs as potential biomarkers for Parkinson's disease.