PXD022662

PXD022662 is an original dataset announced via ProteomeXchange.

Title	Development of a multiplex targeted mass spectrometry assay to quantify levels of LRRK2 phosphorylated Rab substrates and Ser910 and Ser935 biomarker sites
Description	Mutations that increase the protein kinase activity of LRRK2 are one of the most common causes of familial Parkinson's disease. LRRK2 phosphorylates a subset of Rab GTPases within their Switch-II motif, impacting interaction with effectors. We describe and validate a new, multiplex targeted mass spectrometry assay to quantify endogenous levels of LRRK2 phosphorylated Rab substrates (Rab1, Rab3, Rab8, Rab10, Rab35 and Rab43) as well as total levels of Rabs, LRRK2 and phosphorylation of the LRRK2 Ser910 and Ser935 biomarker sites. Exploiting this assay, we quantify for the first time the relative levels of each of the pRab proteins in different cells (mouse embryonic fibroblasts & human neutrophils) and mouse tissues (brain, lung, kidney and spleen). We define how each of the different pRab proteins are impacted by Parkinson’s pathogenic LRRK2[R1441C] and VPS35[D620N] mutations as well as LRRK2 inhibitors. We find that the VPS35[D620N], but not LRRK2[R1441C] mutation, enhances Rab1 phosphorylation in a manner blocked by administration of an LRRK2 inhibitor, providing the first evidence that LRRK2 can phosphorylate Rab1 physiologically. We argue that this targeted mass spectrometry assay can replace immunoblotting approaches currently deployed to assess LRRK2 Rab signalling pathway.
HostingRepository	PRIDE
AnnounceDate	2021-03-17
AnnouncementXML	Submission_2021-03-17_12:47:13.424.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Raja Sekhar Nirujogi
SpeciesList	scientific name: Mus musculus (Mouse); NCBI TaxID: 10090; scientific name: Homo sapiens (Human); NCBI TaxID: 9606;
ModificationList	phosphorylated residue; iodoacetamide derivatized residue; deamidated residue
Instrument	Q Exactive HF

Revision	Datetime	Status	ChangeLog Entry
0	2020-11-23 07:47:51	ID requested
⏵ 1	2021-03-17 12:47:14	announced

Nirujogi RS, Tonelli F, Taylor M, Lis P, Zimprich A, Sammler E, Alessi DR, Development of a multiplexed targeted mass spectrometry assay for LRRK2-phosphorylated Rabs and Ser910/Ser935 biomarker sites. Biochem J, 478(2):299-326(2021) [pubmed]

submitter keyword: Parkinson's disease, LRRK2, RAB-GTPases, PRM, LC-MS/MS

Dario R. Alessi
contact affiliation	Medical Research Council (MRC) Protein Phosphorylation and Ubiquitylation Unit, School of Life Sciences, University of Dundee, Dow Street, Dundee DD1 5EH, UK
contact email	d.r.alessi@dundee.ac.uk
lab head
Raja Sekhar Nirujogi
contact affiliation	MRC Protein Phosphorylation Unit, university of Dundee
contact email	rnirujogi@dundee.ac.uk
dataset submitter

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PRIDE project URI

• PRIDE
  1. PXD022662
     1. Label: PRIDE project
     2. Name: Development of a multiplex targeted mass spectrometry assay to quantify levels of LRRK2 phosphorylated Rab substrates and Ser910 and Ser935 biomarker sites