PXD043498-2

PXD043498 is an original dataset announced via ProteomeXchange.

Title	Puromycin acetyltransferase-mediated cell-specific labeling of de novo synthesized proteins, Part II
Description	Puromycin and its derivative O-propargyl puromycin (OPP) enable detection of nascent proteins. Use of these metabolic labels in complex mixtures of cells leads to indiscriminate tagging of nascent proteomes independent of cell type. Here we have used puromycin N-acetyltransferase (PAT), for cell-specific metabolic labelling with puromycin or OPP. This approach, which we named Puromycin Inactivation for Cell-Selective proteome Labelling (PICSL), is based on efficient inactivation of puromycin or OPP in cells expressing PAT and detection of translation in other cell types. We performed two mass spectrometry experiments using cell-specific expression of PAT, metabolic labeling of cells with OPP and streptavidin pulldown of biotinylated de novo synthesized proteins. In the first proof-of-principle experiment (Part I) we mixed human (HEK293) and mouse (Neuro2a) cells and labeled them with OPP, comparing this with another sample where HEK293 cells expressed PAT. We show specific reduction of human-specific peptides in de novo synthesized proteome in HEK-PAT cells. In the second experiment (Part II), we used a primary co-culture of rat cortical neurons (expressing PAT in some samples) and glia, performed OPP labeling and pulldown of de novo synthesized proteins.
HostingRepository	PRIDE
AnnounceDate	2023-11-14
AnnouncementXML	Submission_2023-11-14_09:11:48.182.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Indrek Koppel
SpeciesList	scientific name: Rattus rattus (Black rat); NCBI TaxID: 10117;
ModificationList	No PTMs are included in the dataset
Instrument	Q Exactive HF

Revision	Datetime	Status	ChangeLog Entry
0	2023-07-03 07:49:07	ID requested
1	2023-10-24 13:30:29	announced
⏵ 2	2023-11-14 09:11:49	announced	2023-11-14: Updated project metadata.
3	2024-10-22 06:12:42	announced	2024-10-22: Updated project metadata.

Cabrera-Cabrera F, Tull H, Capuana R, Kasvandik S, Timmusk T, Koppel I, Cell type-specific labeling of newly synthesized proteins by puromycin inactivation. J Biol Chem, 299(9):105129(2023) [pubmed]

submitter keyword: OPP-ID, shotgun proteomics, streptavidin pulldown, rat cortical neuron-glia co-culture

Indrek Koppel
contact affiliation	Department of Chemistry and Biotechnology Tallinn University of Technology Estonia
contact email	indrek.koppel@taltech.ee
lab head
Indrek Koppel
contact affiliation	Tallinn University of Technology
contact email	indrek.koppel@taltech.ee
dataset submitter

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PRIDE project URI

• PRIDE
  1. PXD043498
     1. Label: PRIDE project
     2. Name: Puromycin acetyltransferase-mediated cell-specific labeling of de novo synthesized proteins, Part II