PXD002111

PXD002111 is an original dataset announced via ProteomeXchange.

Title	Analysis of signaling endosome composition and dynamics using SILAC in embryonic stem cell-derived neurons
Description	Neurons require efficient transport mechanisms such as fast axonal transport to ensure neuronal homeostasis and survival. Neurotrophins and their receptors are conveyed via fast axonal retrograde transport of signaling endosomes to the soma, where they elicit transcriptional responses. Despite the essential roles of signaling endosomes in neuronal differentiation and survival, little is known about their molecular identity, dynamics and regulation. Gaining a better mechanistic understanding of these organelles and their kinetics is crucial, given the growing evidence linking vesicular trafficking deficits to neurodegeneration. Here, we exploited an affinity purification strategy using the binding fragment of tetanus neurotoxin (HCT) conjugated to monocrystalline iron oxide nanoparticles (MIONs), which in motor neurons, is transported in the same carriers as neurotrophins and their receptors. To quantitatively assess the molecular composition of HCT-containing signaling endosomes, we have developed a protocol for triple Stable Isotope Labeling with Amino acids in Cell culture (SILAC) in embryonic stem cell-derived motor neurons. After HCT internalization, retrograde carriers were magnetically isolated at different time points and subjected to mass-spectrometry and gene ontology analyses. This purification strategy is highly specific, as confirmed by the presence of essential regulators of fast axonal transport in the make-up of these organelles. Our results indicate that signaling endosomes undergo a rapid maturation with the acquisition of late endosome markers following a specific time-dependent kinetics. Strikingly, signaling endosomes are specifically enriched in proteins known to be involved in neurodegenerative diseases and neuroinfection. Moreover, we highlighted the presence of novel components, whose precise temporal recruitment on signaling endosomes might be essential for proper sorting and/or transport of these organelles. This study provides the first quantitative proteomic analysis of signaling endosomes isolated from motor neurons and allows the assembly of a functional map of these axonal carriers involved in long-range neuronal signaling.
HostingRepository	PRIDE
AnnounceDate	2015-12-21
AnnouncementXML	Submission_2015-12-21_06:08:37.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Vesela Encheva
SpeciesList	scientific name: Mus musculus (Mouse); NCBI TaxID: 10090;
ModificationList	monohydroxylated residue; acetylated residue; iodoacetamide derivatized residue
Instrument	Q Exactive

Revision	Datetime	Status	ChangeLog Entry
0	2015-04-28 01:43:47	ID requested
⏵ 1	2015-12-21 06:08:38	announced

Debaisieux S, Encheva V, Chakravarty P, Snijders AP, Schiavo G, Analysis of Signaling Endosome Composition and Dynamics Using SILAC in Embryonic Stem Cell-Derived Neurons. Mol Cell Proteomics, 15(2):542-57(2016) [pubmed]

submitter keyword: SILAC, motor neurons, mass spectrometry, signalling endosomes

Ambrosius P. Snijders
contact affiliation	Mass Spectrometry Proteomics and Metabolomics STP The Francis Crick Institute
contact email	Bram.Snijders@crick.ac.uk
lab head
Vesela Encheva
contact affiliation	The Francis Crick Institute
contact email	Vesela.Encheva@crick.ac.uk
dataset submitter

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PRIDE project URI

• PRIDE
  1. PXD002111
     1. Label: PRIDE project
     2. Name: Analysis of signaling endosome composition and dynamics using SILAC in embryonic stem cell-derived neurons