Immunoblotting of macrophages and microglia
By onThis protocol describes the preparation from cell lysate from cultured cells and immunoblotting procedure.
Mitochondrial complex activity assays
By onMitochondria complex activity assays measure the activity levels of the different complexes of the mitochondrial electron transport chain (ETC).
Reconstitution of cargo-induced LC3 lipidation in mammalian selective autophagy
By onCode for analysis of multicolor fluorescence images to quantify fluorescence intensity at the perimeter of giant unilamellar vesicles (GUVs).
Cryo-EM sample preparation
By onThis protocol details how to prepare cryo-EM samples of 3xFLAG-SHIP164Δ901-1099
Midbrain viral injections for striatal fiber photometry in mice
By onThis protocol describes how to: - Inject AAV viruses into the midbrain (SNc: substantia nigra pars compacta, or VTA: ventral tegmental area) of mice - Train mice for head-fixed running on a cylindrical treadmill, and to receive rewards and air puffs while head-fixed - Implant head plates on mice for head-fixation during behavior (optional) While this protocol is focused on injections of AAV viruses into the midbrain, it can be easily modified to inject into other brain regions or other viruses.
Structural basis for ATG9A recruitment to the ULK1 complex in mitophagy initiation
By onHere, the authors examine the structural interaction between ATG9A and components of the ULK1 complex to better understand the process of the PINK1- and Parkin- dependent mitophagy pathway implicated in Parkinson's disease.
Dopamine transporter and synaptic vesicle sorting defects underlie auxilin-associated Parkinson’s disease
By onAuxilin participates in the uncoating of clathrin-coated vesicles (CCVs), thereby facilitating synaptic vesicle (SV) regeneration at presynaptic sites. Auxilin (DNAJC6/PARK19) loss-of-function mutations cause early-onset Parkinson's disease (PD). Here, we utilized auxilin knockout (KO) mice to elucidate the mechanisms through which auxilin deficiency and clathrin-uncoating deficits lead to PD. Auxilin KO mice display cardinal features of PD, including progressive motor deficits, α-synuclein pathology, nigral dopaminergic loss, and neuroinflammation. Significantly, treatment with L-DOPA ameliorated motor deficits. Unbiased proteomic and neurochemical analyses of auxilin KO brains indicated dopamine dyshomeostasis. We validated these findings by demonstrating slower dopamine reuptake kinetics in vivo, an effect associated with dopamine transporter misrouting into axonal membrane deformities in the dorsal striatum. Defective SV protein sorting and elevated synaptic autophagy also contribute to ineffective dopamine sequestration and compartmentalization, ultimately leading to neurodegeneration. This study provides insights into how presynaptic endocytosis deficits lead to dopaminergic vulnerability and pathogenesis of PD.
pCAG-ATG13 (1-197)
By onPlasmid: To make ATG101/ATG13 HORMA complex for expression in mammalian cells.
Gene editing of YIPF4, CALCOCO1, and ATG7 in HeLa cells
By onThe text outlines the process of generating YIPF4, FIP200, and ATG7 knockout cell lines in HeLa cells through CRISPR-Cas9 technology.
Orchestration of selective autophagy by cargo receptors
By onThe authors review recent insights into the mechanisms of action of cargo receptors in selective autophagy by focusing on the roles of sequestosome-like cargo receptors in the degradation of misfolded, ubiquitinated proteins and damaged mitochondria.
Genome-wide screen reveals Rab12 GTPase as a critical activator of Parkinson’s disease-linked LRRK2 kinase
By onThe data support a model in which Rab12 binding to a new site in the LRRK2 Armadillo domain activates LRRK2 kinase for Rab phosphorylation
Endogenous coimmunoprecipitation
By onProtocol used for immunoprecipitation of HSP60 and LONP1 in HEK cells to show the interaction with V5-Flag-tagged WT-GCase
Whole-cell patch-clamping of cultured human neurons
By onThis protocol describes procedures of the whole-cell patch clamping of human neurons derived from induced pluripotent stem cells and cultured in adherent monolayer.
Erythroid Differentiation Dependent Interaction of VPS13A with XK at the Plasma Membrane of K562 Cells
By onMutations in VPS13A and XK lead to Chorea Acanthocytosis and McLeod syndrome, causing neurodegeneration and abnormal red blood cells. VPS13A binds XK at the ER and plasma membrane contacts in differentiated erythroblasts.
Loss of tau expression attenuates neurodegeneration associated with α-synucleinopathy
By onThe authors examine the role of tau in the onset and progression of αS pathology using a transgenic mouse model of α-synucleinopathy lacking mouse tau expression.
Genome-wide Analysis of Motor Progression in Parkinson Disease
By onThe genetic basis of Parkinson disease (PD) motor progression is largely unknown. Previous studies of the genetics of PD progression have included small cohorts and shown a limited overlap with genetic PD risk factors from case-control studies. Here, we have studied genomic variation associated with PD motor severity and early-stage progression in large longitudinal cohorts to help to define the biology of PD progression and potential new drug targets.
Microscopy-based evaluation of Parkin translocation and mitophagy in FBXO7-/- cell lines
By onAssociated with preprint:
Impact of 100 LRRK2 variants linked to Parkinson’s Disease on kinase activity and microtubule binding
By onThis study systematically investigates 100 LRRK2 variants linked to PD, and provides rationale for variant carrier inclusion/exclusion in ongoing and future LRRK2 inhibitor clinical trials.
Primary data associated with doi: 10.1101/2022.04.25.489459 (A Feed-forward Pathway Drives LRRK2 kinase Membrane Recruitment and Apparent Activation)
By onRaw immunoblotting data; tabular data for quantifications; immunofluorescence images.
