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Catalog
ASAP is committed to accelerating the pace of discovery and informing a path to a cure for Parkinson’s disease through collaboration, research-enabling resources, and data sharing. We’ve created this catalog to showcase the research outputs and tools developed by ASAP-funded programs.
Article
Published: PKC isoforms activate LRRK1 kinase by phosphorylating conserved residues (Ser1064, Ser1074, and Thr1075) within the CORB GTPase domain
Published: This study describes how PKC isoforms are able to phosphorylate LRRK1 at three sites in a key regulatory domain of the protein (GTPase domain) inducing LRRK1’s kinase activity. Interestingly, this is not seen with the PD-associated LRRK2, suggesting that PKC isoforms do not regulate LRRK2. View original preprint.
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Article
Global ubiquitylation analysis of mitochondria in primary neurons identifies physiological Parkin targets following activation of PINK1
Published: Mutations in PINK1 and Parkin are implicated in PD via abherrant mitophagy. The authors identified ubiquitylated substrates of endogenous Parkin in mouse neurons by proteomic analysis. They identified and validated 22 protein targets of Parkin that are conserved in human neurons providing a functional Parkin landscape in neuronal cells. View original preprint.
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Article
Global ubiquitylation analysis of mitochondria in primary neurons identifies endogenous Parkin targets following activation of PINK1
Published: Loss-of-function mutations in Parkin cause disruption of mitophagy and are associated with PD. Yet, much of the biology surrounding Parkin function has taken place in artificial cell systems. The authors used human neurons to identify and validate 22 protein targets of Parkin, providing a functional Parkin landscape in neuronal cells.
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Article
PKC isoforms activate LRRK1 kinase by phosphorylating conserved residues (Ser1064, Ser1074, and Thr1075) within the CORB GTPase domain
Leucine-rich-repeat-kinase 1 (LRRK1) and its homologue LRRK2 are multidomain kinases possessing a ROC-CORA-CORB containing GTPase domain and phosphorylate distinct Rab proteins. LRRK1 loss of function mutations cause the bone disorder osteosclerotic metaphyseal dysplasia, whereas LRRK2 missense mutations that enhance kinase activity cause PD. Here, the authors study the mechanism controlling LRRK1 activity and reveal a novel unexpected activation mechanism. View preprint.
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Dataset
Primary data associated with the manuscript “Golgi-IP, a novel tool for multimodal analysis of Golgi molecular content” (doi.org/10.1101/2022.11.22.517583)
Raw data files used for the manuscript “Golgi-IP, a novel tool for multimodal analysis of Golgi molecular content” (doi.org/10.1101/2022.11.22.517583).
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Protocol
CRISPR-Cas9 screen in NIH-3T3 cells to identify modulators of LRRK2 function
This protocol describes a pooled, CRISPR Cas9 screen to identify modulators of LRRK2 activity. The CRISPR-Cas9 based screen is carried out in mouse cells using a ready-to-use pooled guide RNA (gRNA) mouse library consisting of 78,637 gRNAs targeting 19,674 genes and an extra 1,000 control gRNAs.
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Protocol
Untargeted lipidomics analysis for Golgi immunopurification (Golgi-IP)
This protocol provides details for analyzing GolgiIP lipidomics samples using liquid chromatography mass spectrometry (LC-MS) for nonpolar lipid profiling.
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Protocol
Golgi immunopurification (Golgi-IP) for subcellular lipid profiling
This protocol provides details for preparing Golgi-IP lipidomics samples.
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Protocol
Untargeted metabolomics analysis for Golgi immunopurification (Golgi-IP)
This protocol provides details for analyzing GolgiIP metabolomics samples using liquid chromatography mass spectrometry (LC-MS) for polar metabolite profiling.
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Protocol
Golgi immunopurification (Golgi-IP) for subcellular metabolite profiling
This protocol provides details for preparing Golgi-IP metabolomics samples.
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Code
Python script for Golgi cytoscape analysis
Custom Python script used for the cytoscape network analysis reported in doi.org/10.1101/2022.11.22.517583 (Golgi-IP, a novel tool for multimodal analysis of Golgi molecular content).
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Article
Parkinson’s VPS35[D620N] mutation induces LRRK2 mediated lysosomal association of RILPL1 and TMEM55B
Our study uncovers a pathway through which dysfunctional lysosomes resulting from the VPS35[D620N] mutation recruit and activate LRRK2 on the lysosomal surface, driving assembly of the RILPL1-TMEM55B complex.
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Article
Isotope tracing in health and disease
Here, the authors review recent work utilizing metabolic tracing to study health and disease, and highlight its application to interrogate subcellular, intercellular, and in vivo metabolism. The authors further discuss the current challenges and opportunities to expand the utility of isotope tracing to new research areas.
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Protocol
Co-immunoprecipitation protocol to study LRRK2 binding to Rab12 in a cell-based assay
The immunoprecipitation (IP) of FLAG-tagged LRRK2 from whole cell lysates to assess its interaction with Rab12. This method can be used to screen the impact that LRRK2 mutations have on its binding to Rab12 in cells, as well as the effect of any compound or cell treatment on LRRK2 interaction with Rab12.
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