However, the mechanisms by which GBA mutations contribute to the pathogenesis of PD are not completely understood, and many different and non-exclusive hypotheses have been proposed to explain the observed relationship between alterations in GCase and α-syn pathology 6. Both mutations result in misfolded proteins and a significant reduction in GCase activity.

N409S) are the most common GBA mutations and account for 70% of all GBA mutant alleles in PD patients worldwide 5.

Patients with GD and mutant GBA carriers have a fivefold greater lifetime risk of developing PD than that in the general population 4. Biallelic mutations in the GBA gene cause Gaucher’s disease (GD), a lysosomal storage disorder characterized by decreased GCase activity and subsequent accumulation of GlcCer and glucosylsphingosine (GlcSph) in several organs. GBA encodes the lysosomal enzyme β-glucocerebrosidase (GCase), which hydrolyses glucosylceramide (GlcCer) to ceramide and glucose. Among these factors, mutations in the GBA gene are the main genetic risk factor for developing PD 3, 4. The etiology of the disorder involves an interplay of different factors, including aging, genetic susceptibility, and environmental factors. Several lines of evidence indicate that aberrant α-synuclein misfolding, accumulation, and aggregation are the major pathogenic processes in PD 2. The main neuropathological hallmark of PD is the presence of protein inclusions, known as Lewy bodies, composed mainly of aggregated α-synuclein (α-syn) in surviving neurons. Parkinson’s disease (PD) is an age-related neurodegenerative disorder characterized by a progressive loss of dopaminergic neurons in the substantia nigra pars compacta (SNpc), causing striatal dopamine depletion and, as a consequence, a combination of motor symptoms, including resting tremors, bradykinesia, and rigidity 1. The findings of this study highlighted the role of lysosomal function and lipid metabolism in PD and allowed us to describe a molecular mechanism to understand how mutations in GBA can contribute to an abnormal accumulation of different α-synuclein neurotoxic species in PD pathology.

These local events, occurring only at a subcellular level, lead to an impairment of autophagy pathways, particularly chaperone-mediated autophagy, the main α-synuclein degradative pathway. The loss of GCase activity generates extensive lysosomal dysfunction, promoting the loss of activity of other lysosomal enzymes, affecting lysosomal membrane stability, promoting intralysosomal pH changes, and favoring the intralysosomal accumulation of sphingolipids and cholesterol. We performed a deep analysis of the consequences triggered by the presence of mutant GBA protein and the loss of GCase activity in different cellular compartments, focusing primarily on the lysosomal compartment, and analyzed in detail the lysosomal activity, composition, and integrity. In this study, we generated a set of differentiated and stable human dopaminergic cell lines that express the two most prevalent GBA mutations as well as GBA knockout cell lines as a in vitro disease modeling system to study the relationship between mutant GBA and the abnormal accumulation of α-synuclein. Mutations in the GBA gene that encodes the lysosomal enzyme β-glucocerebrosidase (GCase) are a major genetic risk factor for Parkinson’s disease (PD).

Because syn