Protocol Collection: Combinatorial selective ER-phagy remodels the ER during neurogenesis
By Julia Julia Leonard onThis molecular inventory of ER proteome remodeling and versatile genetic toolkit provides a quantitative framework for understanding contributions of individual ER-phagy receptors for reshaping ER during cell state transitions.
Lentivirus preparation for neuronal transdifferentiation
By Julia Julia Leonard onThis is a protocol for the preparation of Lentiviruses.
Human primary fibroblast culture
By Julia Julia Leonard onThis is a protocol for human primary fibroblast culture.
Neuronal transdifferentiation from human primary adult fibroblasts
By Julia Julia Leonard onThis is a protocol for the direct conversion of human primary fibroblasts into neurons using a combination of transcription factor- and small molecule-based approach. The majority of converted neurons showing characteristics of cortical glutamatergic neurons.
Cytokine profiling analysis on conditioned medium of human neurons using Luminex multiplex assay
By Julia Julia Leonard onThis protocol is used to identify secreted inflammatory factors by cytokine profiling of the conditioned medium from human transdifferentiated neurons of healthy donors and AD patients.
Immunohistochemistry on free-floating and paraffin-embedded tissue sections
By Julia Julia Leonard onThis protocol is used for free-floating frozen (30-50 microns) and paraffin-embedded (10 microns) tissue sections.
Proteome of R1441C and G2019S LRRK2 knock in mice after striatal motor learning task
By Julia Julia Leonard onTo identify the signaling pathways that underlie the motor learning deficits, specifically in the RC mice, the authors performed six-plex tandem mass tag (TMT) quantitative mass spectrometry (MS) to compare the protein expression of either RC or GS and WT mice following the five days of the rotarod test.
Tabular data of fast-scanning voltammetry and physiology experiments in LRRK2 KI mice
By Julia Julia Leonard onThe tabular data of fast-scanning cyclic voltammetry and physiology experiments from manuscript: "R1441C and G2019S LRRK2 knockin mice have distinct striatal molecular, physiological, and behavioral alterations." by Xenias et al.
Tabular data of striatal motor learning experiments in LRRK2 KI mice
By Julia Julia Leonard onThe tabular data of the behavioral experiments described in the manuscript titled, "R1441C and G2019S LRRK2 knockin mice have distinct striatal molecular, physiological, and behavioral alterations."
Fast scan cyclic voltammetry
By Julia Julia Leonard onThis protocol describes the ex vivo fast-scan cyclic voltammetry (FSCV) technique for detecting dopamine in the dorsal striatum in mice.
Behavioral tests in rodents coupled with dopamine signaling manipulations
By Julia Julia Leonard onThese two protocols describe behavioral tests associated with Parkinson’s disease phenotypes in mice. The striatal motor learning protocol (rotarod test) is a two-phase paradigm assessing striatal motor learning under reversible dopamine antagonism.
Viral-mediated short-hairpin RNA knockdown
By Julia Julia Leonard onStereotaxic injection of viral vectors for gene knockdown experiments: The AAV vectors for knocking down the gene of interest were custom-made with Vector Biolabs using AAV-GFP-U6 vector for AAV1 packaging (Cat no, 7040, Vector Biolabs). They were then unilaterally injected into the striatum of mice as the protocol details.
Preparation of fresh paraformaldehyde for mouse perfusion and tissue fixation
By Julia Julia Leonard onPreparation of fresh paraformaldehyde for mouse perfusion and tissue fixation.
Unconventional secretion of alpha-synucein mediated by palmitoylated DNAJC5 oligomers
By Julia Julia Leonard onThis protocol, carried out according to the manufacturer’s instructions, uses a classical ELISA experimental design with sensitive electrochemical detection to provide a measure of 𝜶-synuclein concentration in the conditioned media of cultured iPSC-derived human cells.
Mechanism of human PINK1 activation at the TOM complex in a reconstituted system
By Julia Julia Leonard onPreprint: 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.
Tandem Mass Tag (TMT) label-based mass spectrometry analysis of RILPL1 interactors
By Julia Julia Leonard onPrimary mass spectrometry data associated with Fig.5 in the manuscript, "Parkinson’s VPS35 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
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
By Julia Julia Leonard onPrimary mass spectrometry data associated with Fig.1 and Fig.2 in the manuscript "Parkinson’s VPS35 mutation induces LRRK2 mediated lysosomal association of RILPL1 and TMEM55B" (Phospho-Rab and RILPL1 enrichment at VPS35 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.
Detection of accessible cholesterol in primary cilia using purified His-ALOD4-mNeon in 3T3 Fibroblasts
By Julia Julia Leonard onThe 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.
In vivo reduction of age-dependent neuromelanin accumulation mitigates features of Parkinson’s disease
By savannah onCollection of all datasets (Metabolomics, microscopy, and behavioral data) included in the article entitled "In vivo reduction of age-dependent neuromelanin accumulation mitigates features of Parkinson’s disease."
Unique functional responses differentially map onto genetic subtypes of dopamine neurons
By savannah onPublished: Genetic strategies to isolate dopaminergic subtypes lead to the establishment of a novel subtype of dopamine neurons within the mouse substantia nigra. The results show that the neural activity patterns of the genetically identified subtypes map to differing features of locomotion, such as acceleration, deceleration, and appetitive. View original preprint.