PXD009453

PXD009453 is an original dataset announced via ProteomeXchange.

Title	TAILS N-Terminomics and Proteomics Show Complex Regulation of Proteolytic Processing by O-glycosylation
Description	Proteolytic processing is an irreversible post-translational modification functioning as a ubiquitous regulator of cellular activity. Protease activity is tightly regulated by control of gene expression, compartmentalisation of enzyme and substrate, zymogen activation, enzyme inactivation, and substrate availability. Emerging evidence suggests that proteolysis can also be regulated by substrate glycosylation and that glycosylation of individual sites on a substrate can decrease, or in rare cases, increase its sensitivity to proteolysis. In the present study we investigated the relationship between site-specific O-glycosylation and proteolytic cleavage of extracellular proteins. By in silico analysis we found a significant association between O-glycosylation sites and cleavage sites. We then used a positional proteomic strategy, Terminal Amine Isotopic Labelling of Substrates (TAILS) to map in vivo cleavage sites in HepG2 cells with and without one of the key initiating GalNAc-transferases, GalNAc-T2. To reduce complexity, the comparison were performed in so-called SimpleCells. In these cells the Core 1 β3-galactosyltransferase-specific molecular chaperone (COSMC; C1GALT1C1) has been knocked out, resulting in global truncation of O-glycans leaving only the initial GalNAc. Surprisingly, we found that loss of GalNAc-T2 resulted in not only an increase in cleavage, but also a decrease in cleavage across a broad range of other substrates, including key regulators of the protease network. GALNT2 has previously been identified as a candidate gene for dyslipidaemia and has been shown to specifically glycosylate central regulators of lipid homeostasis, here we find altered processing of several of these regulators including Apolipoproteins B (ApoB) and the Phospholipid transfer protein (PLTP), providing new clues to the link between GALNT2 and lipid homeostasis. In contrast to the findings from the majority of previous studies, these results indicate that O-glycosylation can both decrease and increase substrate cleavage. We challenged this system with exogenous matrix metalloproteinase 9 (MMP9) and neutrophil elastase and observed substantial changes to the neutrophil elastase N-terminome, but no evidence for a similar role for MMP9. Overall, we show that loss of O-glycosylation leads to a general decrease in cleavage, that GalNAc-T2 O-glycosylation affects key regulators of the cellular proteolytic network, including multiple members of the Serpin family.
HostingRepository	PRIDE
AnnounceDate	2018-10-08
AnnouncementXML	Submission_2018-10-08_04:35:31.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Sergey Vakhrushev
SpeciesList	scientific name: Homo sapiens (Human); NCBI TaxID: 9606;
ModificationList	O-(N-acetylamino)hexosyl-L-serine; O-(N-acetylamino)hexosyl-L-threonine; monohydroxylated residue; iodoacetamide derivatized residue
Instrument	LTQ Orbitrap Velos; Orbitrap Fusion ETD

Revision	Datetime	Status	ChangeLog Entry
0	2018-04-10 07:02:01	ID requested
⏵ 1	2018-10-08 04:35:32	announced

King SL, Goth CK, Eckhard U, Joshi HJ, Haue AD, Vakhrushev SY, Schjoldager KT, Overall CM, Wandall HH, -glycosylation. J Biol Chem, 293(20):7629-7644(2018) [pubmed]

ProteomeXchange project tag: Biology/Disease-Driven Human Proteome Project (B/D-HPP), Glycoproteomics (B/D-HPP)

curator keyword: Biological

submitter keyword: Human, LC-MSMS, O-glycoproteomics, Degradomics, TAILS

Hans Wandall
contact affiliation	University of Copenahgen, Copenhagen Center for Glycomics
contact email	hhw@sund.ku.dk
lab head
Sergey Vakhrushev
contact affiliation	Department of Cellular and Molecular Medicine
contact email	seva@sund.ku.dk
dataset submitter

Dataset FTP location NOTE: Most web browsers have now discontinued native support for FTP access within the browser window. But you can usually install another FTP app (we recommend FileZilla) and configure your browser to launch the external application when you click on this FTP link. Or otherwise, launch an app that supports FTP (like FileZilla) and use this address: ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2018/10/PXD009453

PRIDE project URI

• PRIDE
  1. PXD009453
     1. Label: PRIDE project
     2. Name: TAILS N-Terminomics and Proteomics Show Complex Regulation of Proteolytic Processing by O-glycosylation