Untargeted lipidomics analysis for Golgi immunopurification (Golgi-IP)
By onThis protocol provides details for analyzing GolgiIP lipidomics samples using liquid chromatography mass spectrometry (LC-MS) for nonpolar lipid profiling.
Expression and purification MBP-ATG9 Constructs
By onExpression and purification from HEK cells of ATG13, ATG101 and FOLDON-ATG9A proteins.
Confirming circRNA expression by qPCR
By onThis protocol delineates a qPCR method to confirm the expression of circRNAs extracted from brain samples.
Glucosylceramidase Beta (GBA) Genotyping
By onThis protocol details the steps for GBA genotyping. This protocol has been adapted from the PRoBaND Clinical Consortium (incorporating methods described by Neuman et al., 2009 and Stone et al., 2000) and has been used for all publications for PRoBaND / Tracking Parkinson's describing clinical data and outcomes with respect to GBA status
Expression and purification Twin-STREP-FLAG tagged ATG13:ATG101 constructs
By onExpression and purification Twin-STREP-FLAG tagged ATG13:ATG101 constructs
PGK EGFP-LC3B
By onPlasmid for mammalian expression of fluorescently-tagged LC3B, marker for autophagic vesicles.
Web portal (shiney ap) for analysis of total proteomes in response to nutrient stress
By onWeb portal (shiney app) for analysis of total proteomes in response to nutrient stress Associated with preprint: https://doi.org/10.1101/2022.12.06.519342
Code for analysis of ER-phagy in iNeurons via proteomics
By onCode for analysis of ER-phagy in iNeurons via proteomics. Associated with preprint: https://doi.org/10.1101/2023.06.26.546565
SoRA microscopy protocol for imaging oligomers in human brain tissue
By onThis protocol gives a step-by-step guide to imaging oligomers in human brain tissue using a spinning disk confocal microscope.
Flow cytometry-based measurement of trafficking of Golgi to the lysosome via autophagy
By onA protocol was developed to measure Golgi trafficking to lysosomes through autophagy in HEK293 cells using Keima-YIPF3/4 reporters with flow cytometry.
RBG Motif Bridge-Like Lipid Transport Proteins: Structure, Functions, and Open Questions
By onThe life of eukaryotic cells requires the transport of lipids between membranes, which are separated by the aqueous environment of the cytosol. Vesicle-mediated traffic along the secretory and endocytic pathways and lipid transfer proteins (LTPs) cooperate in this transport. Until recently, known LTPs were shown to carry one or a few lipids at a time and were thought to mediate transport by shuttle-like mechanisms. Over the last few years, a new family of LTPs has been discovered that is defined by a repeating β-groove (RBG) rod-like structure with a hydrophobic channel running along their entire length. This structure and the localization of these proteins at membrane contact sites suggest a bridge-like mechanism of lipid transport. Mutations in some of these proteins result in neurodegenerative diseases. Here we review the known properties and well-established or putative physiological roles of these proteins, and we highlight the many questions that remain open about their functions.
Sex-specific microglial responses to glucocerebrosidase inhibition
By onMorpho-dynamic analysis occurring in primary cells derived from female and male mice in response to proinflammatory stimulations and glucocerebrosidase inhibition.
Standard Operating Procedure: Mouse Stereotaxic Intracranial Injection Surgery
By onSOP outlines procedures for mouse stereotaxic intracranial injection surgery.
cDNA clone for bacterial expression of human PPM1H D288A
By onPlasmid for bacterial expression of human PPM1H (D288A); contains TEV protease site for removal of 6His tag.
Membrane curvature sensing and stabilization by the autophagic LC3 lipidation machinery
By onDuring autophagy initiation, the curved phagophore is stabilized. Using in vitro reconstitution and MD simulations, authors show that WIPI2 and ATG16L1 bind these curved phagophores and the ATG12-5-16L1 complex is responsible for membrane curvature.
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.
pCAG-MBP-Foldon-ATG9 (723-839)
By onPlasmid for expression of ATG9 C-terminal tail trimer in mammalian cells.