Abstract
A growing number of genes associated with Parkinson’s disease are implicated in the regulation of lysosome function, including LRRK2, whose missense mutations are perhaps the most common monogenic cause of this neurodegenerative disease. These mutations are collectively thought to introduce a pathologic increase in LRRK2 kinase activity, which is currently a major target for therapeutic intervention. Heterozygous carriers of many missense mutations in the GBA1 gene have dramatically increased risk of Parkinson’s disease. A critical question has recently emerged regarding the potential interplay between the proteins encoded by these two disease-linked genes. Our group has recently demonstrated that knockin mutation of a Parkinson’s-linked GBA1 variant induces severe lysosomal and cytokine abnormalities in murine astrocytes and that these deficits were normalized via inhibition of wild-type LRRK2 kinase activity in these cells. Another group independently found that LRRK2 inhibition increases glucocerebrosidase activity in wild-type human iPSC-derived neurons, as well as those whose activity is disrupted by GBA1 or LRRK2 mutation. Fundamental questions remain in terms of the lysosomal abnormalities and the effects of LRRK2 kinase inhibition in human neurons deficient in glucocerebrosidase activity. Here, we further elucidate the physiological crosstalk between LRRK2 signaling and glucocerebrosidase activity in human iPSC-derived neurons. Our studies show that the allelic loss of GBA1 manifests broad defects in lysosomal morphology and function. Furthermore, our data show an increase in both the accumulation and secretion of oligomeric α-synuclein protein in these GBA1-heterozygous-null neurons, compared to isogenic controls. Consistent with recent findings in murine astrocytes, we observed that multiple indices of lysosomal dysfunction in GBA1-deficient human neurons were normalized by LRRK2 kinase inhibition, while some defects were preserved. Our findings demonstrate a selective but functional intersection between glucocerebrosidase dysfunction and LRRK2 signaling in the cell and may have implications in the pathogenesis and treatment of Parkinson’s disease.
Highlights
LRRK2 is a large multi-domain protein that functions both as a kinase and a GTPase (West et al, 2005; Gloeckner et al, 2006; Biosa et al, 2013; Nguyen and Moore, 2017)
We recently showed that a loss-of-function mutation in GBA1 leads to lysosomal defects in murine astrocytes that could be normalized by inhibition of LRRK2 kinase activity (Sanyal et al, 2020)
We used CRISPR/Cas9 based genome editing technology to create isogenic clones of GBA1 heterozygous-null human iPSCs in two independent wildtype healthy control iPSC lines (BR01 and BR33). These cells were first tested for the loss of GCase protein and two clones for each WT iPSC background were chosen for further studies
Summary
LRRK2 is a large multi-domain protein that functions both as a kinase and a GTPase (West et al, 2005; Gloeckner et al, 2006; Biosa et al, 2013; Nguyen and Moore, 2017). Studies in aged LRRK2 knockout rodents and those involving reductions in LRRK2 activity by knockdown or pharmacological interventions have indicated an important role of LRRK2 in maintaining proper lysosomal function (Tong et al, 2010; Herzig et al, 2011; Hinkle et al, 2012). Inhibition of autophagy or endo-lysosomal function leads to an accumulation of αSyn, indicating the importance of this pathway in αSyn degradation (Zimprich et al, 2004; Fornai et al, 2005). ΑSyn proteostasis is fundamentally linked to LRRK2 activity (Cuervo et al, 2004; Fornai et al, 2005; Schapansky et al, 2018). There is an established causal link between altered LRRK2 activity and αSyn metabolism, likely involving dysfunction of the endo-lysosomal system
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