Effect of GBA1 deficiency in human iPSC-derived dopaminergic neurons and astrocytes
Mutations in the gene glucosidase, beta acid 1 (GBA1) are the strongest genetic risk factor for Parkinson's disease (PD) and associate with faster disease progression.  GBA1 is expressed in all cell types of the central nervous system, with some evidence supporting higher expression in glial cells than neurons.  To elucidate possible divergent functions in neurons versus glia, we differentiated human iPSCs generated from an individual with PD heterozygous for the GBA1 pathogenic null variant IVS2+1 (GBA1IVS/+),Mutations in the gene glucosidase, beta acid 1 (GBA1) are the strongest genetic risk factor for Parkinson's disease (PD) and associate with faster disease progression.  GBA1 is expressed in all cell types of the central nervous system, with some evidence supporting higher expression in glial cells than neurons.  To elucidate possible divergent functions in neurons versus glia, we generated human iPSCs from fibroblasts from an individual with PD carrying the GBA1 c.115+1 G>A, also known as IVS2+1 G>A variant (GBA1IVS/+). This intronic GBA1 variant is associated with severe neuronopathic Gaucher's disease and interrupts the canonical splice site following exon 2, leading to nonsense mediated decay and is therefore considered a null allele.  An isogenic GBA1+/+ Rev control was generated by CRISPR/Cas9 genome editing to revert the GBA1 IVS2+1 G>A variant back to wildtype, and in the genome editing process, an iPSC clone homozygous for GBA1 IVS2+1 G>A was also generated (GBA1IVS/IVS). A matched control iPSC clone that remained unedited after CRISPR/Cas9 genome editing, maintaining heterozygosity for GBA1 IVS2+1 G>A, was also isolated (GBA1IVS/+ A4).Transcriptomic analysis of GBA1-deficient neurons and astrocytes revealed cell-type specific profiles. GBA1deficiency in neurons downregulated genes associated with immune response and upregulation of cholesterol synthesis, while GBA1 deficiency in astrocytes downregulated genes associated with translation and upregulated genes involved in extracellular matrix biogenesis, suggestive of gliosis. The transcriptomic profile of GBA1-deficient astrocytes was consistent with neurotoxic reactive astrocytes. Together, these findings indicate that GBA1deficiency has cell type-specific effects, where neurons have increased vulnerability to alterations in endolysosomal trafficking leading to a-synucleinopathy while GBA1 deficiency in astrocytes leads to increased neurotoxic reactivity but not impairment in endolysosomal trafficking and development of a-synuclein pathology. Overall design: RNAseq profiling of dopaminergic neurons and astrocytes differentiatied from 3 iPSC clones generated from an individual with Parkinson's disease heterozygous for the GBA1 c.115+1G>A (IVS2+1), 1 clone homozygous for GBA1 IVS2+1 that is isogenic to the hereozygous GBA1 IVS2+1 clones, 1 clone with wildtype GBA1 that is isogenic to the heterozygous IVS2+1 clones, 1 clone isogenic to the heterozygous IVS2+1 clones that underwent the same CRISPR/Cas9 gene editing process that resulted in the homozygous GBA1 IVS2+1 clone and the GBA1 WT clone but was not edited by CRISPR/Cas9, and 3 iPSC clones generated from a healthy age- and sex-matched individual unrelated to the individual with Parkinson's disease heterozygous for GBA1 IVS2+1.