The effects of the Parkinson’s disease-associated proteins LRRK2 and TMEM175 on the lysosomal profile of cultured mammalian cells under lysosomal stress conditions

Lysosomal dysfunction is increasingly recognised as a central factor in the development of Parkinson’s disease (PD). At least 11 out of the 24 PD-associated genes have been identified by genome-wide association studies (GWAS) to be involved in autophagy-lysosome pathways. Leucine-rich repeat kinase 2 (LRRK2) has been found as the most common genetic risk factor for PD. LRRK2 is involved in various cellular processes, including cell signalling, vesicle trafficking, and the maintenance of lysosomal homeostasis. Another important PD-associated gene encodes the transmembrane protein 175 (TMEM175), a lysosomal cation channel essential for maintaining lysosomal pH and regulating other aspects of endolysosomal function. The mechanisms by which the mutations in LRRK2 or TMEM175 cause PD have not yet been established, which hinders the development of disease-modifying treatments for the cases caused by these mutations. The objective of this study was to investigate differences in lysosomal characteristics in mammalian A549 cells with distinct LRRK2 genotypes, and in human and mouse microglial lines under basal conditions and lysosomal stress. The pathogenic PD-associated LRRK2 mutations have been shown to increase LRRK2 kinase activity, causing increased phosphorylation of its Rab GTPase substrates. This study shows that A549 cells with PD-associated Y1699C LRRK2 mutation were more abundant in phosphorylated Rab10 compared to WT cells. Currently, LRRK2 is considered the major kinase confirmed to phosphorylate Rab10 at T73. However, this work reports increased levels of pRab10 also in LRRK2 KO cells, suggesting that the phosphorylation of Rab10 at the T73 site may be partly independent of LRRK2 activity. This study shows that LRRK2 has a role in the composition and glycosylation patterns of lysosome associated membrane proteins 1 (LAMP1) and 2 (LAMP2) in A549 cells, as well as in their response to lysosomal stress induced by the lysosome-disrupting agents, chloroquine and LLOME. It was found that A549 cells with different LRRK2 genotypes express distinct levels of glycosylated LAMP1 and LAMP2 proteins. Lysosomal stress induced by chloroquine and LLOME was observed to modulate the size and localisation of LAMP1 and LAMP2 depending on LRRK2 genotype. Additionally, preliminary data revealed 10-fold differences in the relative LAMP2 expression between mouse and human microglia cells. The results of this study support the importance of LRRK2 in regulating lysosomal function, showing that PD-associated mutations in LRRK2 may cause alterations in the lysosomal function of the cells. More effective methods are needed to study the role of TMEM175 in the lysosomal profile and the development of PD.

Grantor dc:publisher

Helsingin yliopisto

Year dc:date.issued

2025

Author dc:creator

• Himanka, Ella

• Parkinsonin tauti
• lysosomaalinen stressi
• LRRK2
• TMEM175
• klorokiini
• LLOME
• lysosomin kalvoproteiinit

Statement dc:rights

• In Copyright 1.0

Language dc:language.iso

eng

Handle dc:identifier.uri

http://hdl.handle.net/10138/601463

OAI identifier oai:identifier

oai:helda.helsinki.fi:10138/601463