Evaluation of an adapted semi-automated DNA extraction for human salivary shotgun metagenomics
By onThis study demonstrates that the authors’ semi-automated protocol is suitable for shotgun metagenomic analysis.
Microbes and Parkinson’s Disease: from associations to mechanisms
By onSeveral microbes, including viruses, bacteria, and fungi, have been associated with an increased risk of PD in humans. Microbial infections can induce similar common pathways that are associated with PD, including systemic inflammatory responses, α-synuclein misfolding, and disruption of mitochondria. PD-associated gene mutations can impact host–microbe interactions, suggesting that even familial forms of PD may be influenced by microbes.
Sphingolipid changes in Parkinson L444P GBAmutation fibroblasts promote α-synuclein aggregation
By onIntraneuronal accumulation of aggregated α-synuclein is a pathological hallmark of Parkinson’s disease. Therefore, mechanisms capable of promoting α-synuclein deposition bear important pathogenetic implications. Mutations of the glucocerebrosidase 1 (GBA) gene represent a prevalent Parkinson’s disease risk factor. They are associated with loss of activity of a key enzyme involved in lipid metabolism, glucocerebrosidase, supporting a mechanistic relationship between abnormal α-synuclein–lipid interactions and the development of Parkinson pathology. In this study, the lipid membrane composition of fibroblasts isolated from control subjects, patients with idiopathic Parkinson’s disease and Parkinson's disease patients carrying the L444P GBA mutation (PD-GBA) was assayed using shotgun lipidomics. The lipid profile of PD-GBA fibroblasts differed significantly from that of control and idiopathic Parkinson’s disease cells. It was characterized by an overall increase in sphingolipid levels. It also featured a significant increase in the proportion of ceramide, sphingomyelin and hexosylceramide molecules with shorter chain length and a decrease in the percentage of longer-chain sphingolipids. The extent of this shift was correlated to the degree of reduction of fibroblast glucocerebrosidase activity. Lipid extracts from control and PD-GBA fibroblasts were added to recombinant α-synuclein solutions. The kinetics of α-synuclein aggregation were significantly accelerated after addition of PD-GBA extracts as compared to control samples. Amyloid fibrils collected at the end of these incubations contained lipids, indicating α-synuclein–lipid co-assembly. Lipids extracted from α-synuclein fibrils were also analysed by shotgun lipidomics. Data revealed that the lipid content of these fibrils was significantly enriched by shorter-chain sphingolipids. In a final set of experiments, control and PD-GBA fibroblasts were incubated in the presence of the small molecule chaperone ambroxol. This treatment restored glucocerebrosidase activity and sphingolipid levels and composition of PD-GBA cells. It also reversed the pro-aggregation effect that lipid extracts from PD-GBA fibroblasts had on α-synuclein. Taken together, the findings of this study indicate that the L444P GBA mutation and consequent enzymatic loss are associated with a distinctly altered membrane lipid profile that provides a biological fingerprint of this mutation in Parkinson fibroblasts. This altered lipid profile could also be an indicator of increased risk for α-synuclein aggregate pathology.
x4 GBA Plasmids
By onThe below plasmids are deposited and available via Addgene:https://www.addgene.org/Anthony_Schapira/ . These have been used in publication: 10.1093/hmg/ddac233 188580 WT GBA pcDNA3.1 GBA (Homo sapiens) 188581 E326K GBA pcDNA3.1 GBA (Homo sapiens) 188582 L444P GBA pcDNA3.1 GBA (Homo sapiens) 188583 N370S GBA pcDNA3.1 GBA (Homo sapiens)
Post mortem human substantial nigra TH staining
By onThis protocol is standardized for postmortem (frozen) SN tissue from pathologically diagnosed PD patients and control individuals for Immunofluorescence staining.
Phenotypic effect of GBA1 variants in individuals with and without Parkinson disease: the RAPSODI study
By onThe authors’ results support previous evidence that GBA1-positive PD has a specific phenotype with more severe non-motor symptoms. The authors did not reproduce previous findings of more frequent prodromal PD signs in non-affected GBA1 carriers.
Unilateral intranigral AAV alpha synuclein mouse model
By onAs some protocols have not been made public and are only shared within the network, you may need to login to the protocols.io to view the link.Don’t have a login? Use this special invite link and click join to get started. Click here for detailed instructions on how to set up your free account and view all protocols.
Intracellular Cytokine (ICS) Staining Protocol
By onThis protocol details about intracellular cytokine (ICS) staining.
Subcellular and regional localization of mRNA translation in midbrain dopamine neurons–DropSeqPipeline8
By onThis is a data processing pipeline for analyzing microwell- or DropSeq-like scRNA-seq data. It can also be used for analyzing pooled plate-based scRNA-seq
Transcriptional analysis of peripheral memory T cells reveals Parkinson’s disease-specific gene signatures–Fluorospot SFC
By onFluorospot SFC (combined IFNg, IL-5, and IL-10) data and corresponding HC-PD group IDs described in "Transcriptional analysis of peripheral memory T cells reveals Parkinson’s disease-specific gene signatures"
Differentiation NPCs to Dopaminergic/Midbrain Neurons
By onThis protocol details methods for differentiation of NPCs to Dopaminergic/Midbrain Neurons. This protocol is part of a Collection of protocols (dx.doi.org/10.17504/protocols.io.8epv593dng1b/v1) for the paper "Glucocerebrosidase, a Parkinson´s disease-associated protein, is imported into mitochondria and regulates complex I assembly and function" (https://doi.org/10.21203/rs.3.rs-1521848/v1)
A Single-Cell Atlas of Cell Types, States, and Other Transcriptional Patterns from Nine Regions of the Adult Mouse Brain
By onWe report 690K single-cell transcriptomes from nine different brain regions from adult mice (Frontal and Posterior Cortex, Hippocampus, Thalamus, Cerebellum, Striatum, Globus Pallidus externus/Nucleus Basalis, Entopeduncular / Subthalamic Nuclei, & Substantia Nigra / Ventral Tegmental Area).
Gcase co-immunoprecipitation
By onThe authors developed this protocol to identify protein-protein interactions between the enzyme glucocerebrosidase (GCase) and other proteins in human iPSC-derived Neural Precursor Cells.
Axonal and somatodendritic proteomes of dopamine neurons in the mouse brain
By onDopamine (DA) neurons modulate neural circuits and behaviors via dopamine release from expansive, long range axonal projections. The elaborate cytoarchitecture of DA neurons is embedded within complex brain tissues, making it difficult to access the DA neuronal proteome using conventional methods. Here, we demonstrate APEX2 proximity labeling within genetically targeted neurons in the mouse brain, enabling subcellular proteomics with cell type-specificity. By combining APEX2 biotinylation with mass spectrometry, we mapped the somatodendritic and axonal proteomes of DA neurons. Our dataset reveals the proteomic architecture underlying axonal transport, dopamine transmission, and axonal metabolism in DA neurons. We find a significant enrichment of proteins encoded by Parkinson’s disease-linked genes in dopaminergic axons, including proteins with previously undescribed axonal localization. Our proteomic datasets comprise a significant resource for axonal and DA neuronal cell biology, while the methodology developed here will enable future studies of other neural cell types. This mass spectrometry proteomics dataset is a part of "Subcellular proteomics of dopamine neurons in the mouse brain" (Hobson et. al, 2022)
Flag co-immunoprecipitation
By onProtocol used to pull down V5-FLAG-Gcase interacting proteins in HEK cells
Dopamine transporter and synaptic vesicle sorting defects underlie auxilin-associated Parkinson’s disease
By onAuxilin participates in clathrin uncoating to facilitate presynaptic endocytosis. Loss-of-function mutations of auxilin (PARK19) cause Parkinson’s disease. Using auxilin KO mice, Vidyadhara et al. (2023) show that synaptic vesicle sorting deficits, cytoplasmic dopamine accumulation, dopamine transporter mistrafficking, and synaptic autophagic overload may lead to pathogenesis of Parkinson’s disease in PARK19 patients. This file contains the data set used to generate all the main figures.
A Markov random field model-based approach for differentially expressed gene detection from single-cell RNA-seq data
By onSingle-cell RNA-sequencing technology enables the identification of cell-type-specific differential gene expressions. MARBLES, a new statistical model, effectively detects DE genes across conditions.
Peripheral neuronal activation shapes the microbiome and alters gut physiology
By onThe authors specifically activate ChAT- or TH-expressing gut-associated neurons in mice and perform multi-omics, finding that subsets of peripherally-activated neurons differentially regulate the gut microbiome and host GI physiology.
