LRRK2 RCKW Protein Purification
By onProtein purification protocol for tag-less LRRK2RCKW as done by Leschziner and Reck-Peterson Labs. Same protocol can be used to purify LRRK1RCKW as well. Original Protocol by David Snead. Modified by Yu Xuan Lin and Mariusz Matyszewski for publication.
The Parkinson’s disease protein alpha-synuclein is a modulator of processing bodies and mRNA stability
By onThe paper describes the ability of alpha-synuclein to bind to P-bodies, machinery that regulates the expression of our genes through mRNAs. When alpha-synuclein abnormally accumulates, the physiologic structure and functions of the P-body are lost.
Lentivirus production for primary neuron transduction
By onThis protocol is associated with the following preprint, published on February 19th 2022: The AAA+ chaperone VCP disaggregates Tau fibrils and generates aggregate seeds Itika Saha, Patricia Yuste-Checa, Miguel Da Silva Padilha, Qiang Guo, Roman Körner, Hauke Holthusen, Victoria A. Trinkaus, Irina Dudanova, Rubén Fernández-Busnadiego, Wolfgang Baumeister, David W. Sanders, Saurabh Gautam, Marc I. Diamond, F. Ulrich Hartl, Mark S. Hipp bioRxiv 2022.02.18.481043; doi: https://doi.org/10.1101/2022.02.18.481043
Electron microscope sample preparation technique_V2
By onEM sample preparation protocol that is designed for processing cultured cell sample.
Primary data associated with the manuscript “Genome wide screen reveals Rab12 GTPase as a critical activator of pathogenic LRRK2 kinase” (doi: 10.1101/2023.02.17.529028)
By onPrimary data associated with the manuscript "Genome wide screen reveals Rab12 GTPase as a critical activator of pathogenic LRRK2 kinase"
Combining biomarkers for prognostic modelling of Parkinson’s disease
By onParkinson's disease progression varies among patients. Predicting progression accurately is crucial for clinical trial selection. Blood biomarkers like serum NfL, along with genetic factors (GBA, APOE) can enhance prediction beyond age and phenotype.
Fecal metagenomic sequencing data for PD patients and controls from Rush University Medical Center
By onFecal metagenomic sequencing data associated with Boktor et al. (2023). This dataset includes samples from the Rush University Medical Center cohort.
DQ-BSA assay
By onThis protocol describes the DQ BSA assay for measuring the lysosomal proteolytic activity.
H9 ES AAVS1-NGN2 GRN-/-
By onH9 ES cells modified using CRISPR/Cas9 lack GRN gene and express NEUROG2 in AAVS1 safe harbor locus. Derived from human blastocyst, these embryonic stem cells can differentiate into iNeurons.
Plasmid-reprogramming of human fibroblasts
By onThis protocol is part of a collection of protocols for the paper, "Glucocerebrosidase, a Parkinson's disease-associated protein, is imported into mitochondria and regulates complex I assembly and function"
Quantitative proteomics reveals the selectivity of ubiquitin-binding autophagy receptors in the turnover of damaged lysosomes by lysophagy
By onThe authors used proteomics to develop a quantitative snapshot of the proteins involved in lysophagy. Among the proteins identified, they found that TAX1BP1 and TBK1 are both required for lysophagy.
Gavage – food supplementation for stalled weight gain in mice
By onFeeding protocol for Ndufs2fl/fl mice,
Cell culture, transfection, and imaging
By onThis protocol describes general procedures for culturing HeLa cells, transient transfection, and imaging using an Andor Dragonfly spinning disk confocal system.
L1 retrotransposons drive human neuronal transcriptome complexity and functional diversification
By onThe genetic mechanisms underlying the expansion in size and complexity of the human brain remains poorly understood. L1 retrotransposons are a source of divergent genetic information in hominoid genomes, but their importance in physiological functions and their contribution to human brain evolution is largely unknown. Using multi-omic profiling we here demonstrate that L1-promoters are dynamically active in the developing and adult human brain. L1s generate hundreds of developmentally regulated and cell-type specific transcripts, many which are co-opted as chimeric transcripts or regulatory RNAs. One L1-derived lncRNA, LINC01876, is a human-specific transcript expressed exclusively during brain development. CRISPRi-silencing of LINC01876 results in reduced size of cerebral organoids and premature differentiation of neural progenitors, implicating L1s in human-specific developmental processes. In summary, our results demonstrate that L1-derived transcripts provide a previously undescribed layer of primate- and human-specific transcriptome complexity that contributes to the functional diversification of the human brain.
Lentivirus plasmid: pLV[Exp]-U6>KANSL1_P1_Seq1-hPGK>mApple
By onPlasmid: Plasmid vector encoding a sgRNA sequence which targets the KANSL1 promoter from CRISPRi, under a U6 promoter and a mApple fluorescent reporter. Generated by Vectorbuilder in the pLV backbone.
LRRK2 kinase activity regulates GCase level and enzymatic activity differently depending on cell type in Parkinson’s disease
By onLeucine-rich repeat kinase 2 (LRRK2) is a kinase involved in different cellular functions, including autophagy, endolysosomal pathways, and immune function. Mutations in LRRK2 cause autosomal-dominant forms of Parkinson’s disease (PD). Heterozygous mutations in GBA1, the gene encoding the lysosomal enzyme glucocerebrosidase (GCase), are the most common genetic risk factors for PD. Moreover, GCase function is altered in idiopathic PD and in other genetic forms of the disease. Recent work suggests that LRRK2 kinase activity can regulate GCase function. However, both a positive and a negative correlation have been described. To gain insights into the impact of LRRK2 on GCase, we performed a comprehensive analysis of GCase levels and activity in complementary LRRK2 models, including (i) LRRK2 G2019S knock in (GSKI) mice, (ii) peripheral blood mononuclear cell (PBMCs), plasma, and fibroblasts from PD patients carrying LRRK2 G2019S mutation, (iii) patient iPSCs-derived neurons; (iv) endogenous and overexpressed cell models. In some of these models we found a positive correlation between the activities of LRRK2 and GCase, which was further confirmed in cell lines with genetic and pharmacological manipulation of LRRK2 kinase activity. GCase protein level is reduced in GSKI brain tissues and in G2019S iPSCs-derived neurons, but increased in fibroblasts and PBMCs from patients, suggesting cell-type-specific effects. Overall, our study indicates that LRRK2 kinase activity affects both the levels and the catalytic activity of GCase in a cell-type-specific manner, with important implications in the context of therapeutic application of LRRK2 inhibitors in GBA1-linked and idiopathic PD.
