Cell Type-Specific Transcriptomics Reveals that Mutant Huntingtin Leads to Mitochondrial RNA Release and Neuronal Innate Immune Activation.

The mechanisms by which mutant huntingtin (mHTT) leads to neuronal cell death in Huntington's disease (HD) are not fully understood. To gain new molecular insights, we used single nuclear RNA sequencing (snRNA-seq) and translating ribosome affinity purification (TRAP) to conduct transcriptomic analyses of caudate/putamen (striatal) cell type-specific gene expression changes in human HD and mouse models of HD. In striatal spiny projection neurons, the most vulnerable cell type in HD, we observe a release of mitochondrial RNA (mtRNA) (a potent mitochondrial-derived innate immunogen) and a concomitant upregulation of innate immune signaling in spiny projection neurons. Further, we observe that the released mtRNAs can directly bind to the innate immune sensor protein kinase R (PKR). We highlight the importance of studying cell type-specific gene expression dysregulation in HD pathogenesis and reveal that the activation of innate immune signaling in the most vulnerable HD neurons provides a novel framework to understand the basis of mHTT toxicity and raises new therapeutic opportunities.

LRRK2 Kinase Inhibition Rescues Deficits in Lysosome Function Due to Heterozygous GBA1 Expression in Human iPSC-Derived Neurons.

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.

Lysosome and Inflammatory Defects in GBA1-Mutant Astrocytes Are Normalized by LRRK2 Inhibition.

BACKGROUND: Autosomal recessive mutations in the glucocerebrosidase gene, Beta-glucocerebrosidase 1 (GBA1), cause the lysosomal storage disorder Gaucher's disease. Heterozygous carriers of most GBA1 mutations have dramatically increased Parkinson's disease (PD) risk, but the mechanisms and cells affected remain unknown. Glucocerebrosidase expression is relatively enriched in astrocytes, yet the impact of its mutation in these cells has not yet been addressed. OBJECTIVES: Emerging data supporting non-cell-autonomous mechanisms driving PD pathogenesis inspired the first characterization of GBA1-mutant astrocytes. In addition, we asked whether LRRK2, likewise linked to PD and enriched in astrocytes, intersected with GBA1 phenotypes. METHODS: Using heterozygous and homozygous GBA1 D409V knockin mouse astrocytes, we conducted rigorous biochemical and image-based analyses of lysosomal function and morphology. We also examined basal and evoked cytokine response at the transcriptional and secretory levels. RESULTS: The D409V knockin astrocytes manifested broad deficits in lysosomal morphology and function, as expected. This, however, is the first study to show dramatic defects in basal and TLR4-dependent cytokine production. Albeit to different extents, both the lysosomal dysfunction and inflammatory responses were normalized by inhibition of LRRK2 kinase activity, suggesting functional intracellular crosstalk between glucocerebrosidase and LRRK2 activities in astrocytes. CONCLUSIONS: These data demonstrate novel pathologic effects of a GBA1 mutation on inflammatory responses in astrocytes, indicating the likelihood of broader immunologic changes in GBA-PD patients. Our findings support the involvement of non-cell-autonomous mechanisms contributing to the pathogenesis of GBA1-linked PD and identify new opportunities to correct these changes with pharmacological intervention. © 2020 International Parkinson and Movement Disorder Society.