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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.
Primary data associated with the manuscript 10.1126/science.adi9926 (“Rab29-dependent asymmetrical activation of leucine-rich repeat kinase 2”)
Primary data associated with the manuscript “Rab29-dependent asymmetrical activation of leucine-rich repeat kinase 2” (Hanwen Zhu, Francesca Tonelli, Martin Turk, Alan Prescott, Dario R. Alessi, Ji Sun). These include immunoblotting data and their quantitation, representative confocal microscopy images, and quantitation and statistical analysis of confocal microscopy images.
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Mechanism of human PINK1 activation at the TOM complex in a reconstituted system
Published: The authors demonstrate an essential role of the pore-containing subunit TOM40 and its structurally associated subunits TOM7 and TOM22 for PINK1 activation. These molecular findings will aid in the development of small molecule activators of PINK1 as a therapeutic strategy for PD. View original preprint.
Live cell imaging of yeast cells expressing human PINK1-GFP and the TOM complex subunits
A protocol for live-cell imaging of yeast cells expressing human PINK1 with GFP tag at the C-terminus as well as all the human TOM complex subunits (TOMs 5, 6, 7, 20, 22, 40 and 70).
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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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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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RAB32 Ser71Arg in autosomal dominant Parkinson’s disease: linkage, association, and functional analyses
RAB GTPases are regulators and substrates of LRRK2, and variants in the LRRK2 gene are important risk factors for Parkinson’s disease. Here, the authors explore genetic variability in RAB GTPases within cases of familial Parkinson’s disease.
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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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A computational pipeline to quantify primary cilia in mouse embryonic fibroblasts with CellProfiler
This protocol works with .czi format images which are acquired using a Zeiss laser scanning confocal microscope and are maximum intensity projected.
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Tandem Mass Tag (TMT) label-based mass spectrometry analysis of RILPL1 interactors
Primary mass spectrometry data associated with Fig.5 in the manuscript, “Parkinson’s VPS35[D620N] mutation induces LRRK2 mediated lysosomal association of RILPL1 and TMEM55B” (Association of RILPL1 with a lysosomal membrane integral protein, TMEM55B mediated by LRRK2 activity). Curtain link for the dataset: https://curtain.proteo.info/#/4f15d6c8-9192-4bb2-a2ff-da7aa542025c
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Mass spectrometry analysis of proteome changes in VPS35-WT vs VPS35[D620N] MEFs and VPS35[D620N] MEFs vs LRRK2 inhibitor in whole cell lysates and lysosomes
Primary mass spectrometry data associated with Fig.1 and Fig.2 in the manuscript “Parkinson’s VPS35[D620N] mutation induces LRRK2 mediated lysosomal association of RILPL1 and TMEM55B” (Phospho-Rab and RILPL1 enrichment at VPS35[D620N] mutant lysosome and Lysosomal recruitment of RILPL1 is dependent on LRRK2 kinase activity). Curtain links for the datasets: https://curtain.proteo.info/#/0e673d58-d8f2-4368-996b-0869d5513d46; https://curtain.proteo.info/#/0e673d58-d8f2-4368-996b-0869d5513d46; https://curtain.proteo.info/#/d864df78-e2a5-4a64-99fb-8c5d8b2e1ab8; https://curtain.proteo.info/#/062e0a64-1fd3-4cc1-833a-a5b007d95a3c.
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Detection of accessible cholesterol in primary cilia using purified His-ALOD4-mNeon in 3T3 Fibroblasts
The protocol described here is based upon previously established methods (Kinnebrew et al., 2019; Johnson and Radhakrishnan, 2021) and has also been used successfully to label cilia in RPE cells.
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Primary data associated with the manuscript 10.1073/pnas.2315171120 (“Localization of PPM1H phosphatase tunes Parkinson’s disease-linked LRRK2 kinase-mediated Rab GTPase phosphorylation and ciliogenesis”)
Primary data associated with the manuscript 10.1073/pnas.2315171120 (“Localization of PPM1H phosphatase tunes Parkinson’s disease-linked LRRK2 kinase-mediated Rab GTPase phosphorylation and ciliogenesis”).
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Transfection by electroporation of GFP-LRRK2 and Immunofluorescent imaging of MEFs VPS35 (D620N) mutants stably expressing LysoTag
The authors investigated the colocalization of GFP-LRRK2 with a lysosomal localized TMEM192-3xHA.
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Generation of A549 PPM1H BromoTag CRISPR/CAS9 knock-in cell line
This protocol details the generation of A549 PPM1H BromoTag CRISPR/CAS9 knock-in cell line.
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