Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Discovery of a pathway for endogenous mucin glycodomain catabolism in mammals

ABSTRACT: Mucins are functionally implicated in a range of human pathologies, including cystic fibrosis, influenza, bacterial endocarditis, gut dysbiosis, and cancer. These observations have motivated the study of mucin biosynthesis as well as development of strategies for inhibition of mucin glycosylation. Mammalian pathways for mucin catabolism, however, have remained underexplored. The canonical view, derived from analysis of N-glycoconjugates in human lysosomal storage disorders, is that proteolysis and glycan degradation occur largely independently. Here, we challenge this view by providing genetic and biochemical evidence supporting mammalian proteolysis of heavily O-glycosylated mucin domains, without prior deglycosylation. Using activity screening coupled with mass spectrometry we ascribed mucin-degrading activity in murine liver to the lysosomal protease cathepsin D. Knockout of cathepsin D in a murine model of the human lysosomal storage disorder neuronal ceroid lipofuscinosis resulted in accumulation of mucins in liver-resident macrophages. Our findings suggest that mucin-degrading activity is not limited to the bacterial kingdom and is a component of glycoprotein catabolism in mammalian tissues.

INSTRUMENT(S): Orbitrap Fusion

ORGANISM(S): Homo Sapiens (human) Mus Musculus (mouse)

TISSUE(S): Brain, Liver

SUBMITTER: Nicholas Riley

LAB HEAD: Carolyn Bertozzi

PROVIDER: PXD027616 | Pride | 2022-09-03

REPOSITORIES: Pride

ACCESS DATA

Json Xml

Dataset's files

Source:

			Action	DRS
	2020_05_16_KP_BC1.raw	Raw
	2020_05_16_KP_BC10.raw	Raw
	2020_05_16_KP_BC11.raw	Raw
	2020_05_16_KP_BC12.raw	Raw
	2020_05_16_KP_BC2.raw	Raw

Items per page:

1 - 5 of 50

Similar Datasets

Cell-type specific tagging of glycoproteins in murine-human co-culture samples glycoproteomics lysates

Project description:We report a strategy termed Bio-Orthogonal Cell line-specific Tagging of Glycoproteins (BOCTAG). Cells are equipped by transfection with an artificial biosynthetic pathway (mut-AGX1/NahK/Nh-GalNAc-T2) that transforms an alkyne tagged sugar into the corresponding nucleotide-sugar. Only transfected cells incorporate the bioorthogonal sugar into glycoproteins in the presence of non-transfected cells. The incorporation of alkyne-sugar in glycoproteins allows the installation, by copper(I)-catalyzed reaction, of an acid cleavable biotinylated tag. Glycoproteins are enriched using Streptavidin and. after on-bead digestion, glycopeptides are eluted in acid-conditions to be then analysed by mass spectrometry.

2022-09-10 | PXD035449 | Pride

Cell-type specific tagging of glycoproteins in murine-human co-culture secretome glycoproteomics

2022-09-10 | PXD035445 | Pride

Proteoglycan sequencing - Deep sequencing of complex proteoglycans: a novel strategy for high coverage and site-specific identification of glycosaminoglycan-linked peptides

Project description:Proteoglycans are distributed in all animal tissues and play critical, multifaceted, physiological roles. Expressed in a spatially- and temporally-regulated manner, these molecules regulate interactions among growth factors and cell surface receptors and play key roles in basement membranes and other extracellular matrices. Due to the high degree of glycosylation by glycosaminoglycan (GAG), N-glycan and mucin-type O-glycan classes, the peptide sequence coverage of complex proteoglycans is revealed poorly by standard mass spectrometry-based proteomics methods. As a result, there is little information concerning how proteoglycan site specific glycosylation changes during normal and pathological processes. Here, we developed a workflow to improve sequence coverage and identification of glycosylated peptides in proteoglycans. We applied this workflow to the small leucine-rich proteoglycan decorin and the hyalectan proteoglycans; neurocan, brevican, and aggrecan.

2018-05-22 | PXD008855 | Pride

Glycan shield of the ebolavirus envelope glycoprotein GP

Project description:The envelope glycoprotein GP of the ebolaviruses is essential for host cell attachment and entry. It is also the primary target of the protective and neutralizing antibody response in both natural infection and vaccination. GP is heavily glycosylated with up to 17 predicted N-linked sites, numerous O-linked glycans in its disordered mucin-like domain (MLD), and three predicted C-linked mannosylation sites. Glycosylation of GP is important for host cell attachment to cell-surface lectins, as well as GP stability and fusion activity. Moreover, it has been shown to shield GP from neutralizing activity of serum antibodies. Here, we use mass spectrometry-based glycoproteomics to profile the site-specific glycosylation patterns of ebolavirus GP, including N-, O-, and C-linked glycans.

2022-08-02 | PXD031459 | Pride

O-linked glicoproteomics of gastric cancer cell lines OCUM-1, N87 and KATO-III

Project description:Gastric cancer (GC) is the third leading cause of cancer-related death worldwide, continuously requiring innovative target therapies. The sialyl-lewis A (SLeA) antigen offers tremendous potential for precise cancer targeting. This glycan is rarely expressed by healthy tissues and blood cells but is often present in cancer cells of high metastatic potential as well as in the metastasis. To improve further on the gastric cancer nature of this glycan we have zoomed in on the SLeA-glycoproteome envisaging the identification of possible targetable biomarkers. SLeA-glycoproteins were isolated by immunoprecipitation from well-known cell models of gastric cancer reflecting different histological subtypes of the disease (KATO-III, OCUM-1 and N87 cells). Glycoproteins with affinity for E-selectin were also isolated and analyzed. Proteomics study employed a typical nanoLC-ESI-Orbitrap-MS/MS platform. The well-known molecular heterogeneity of these cell lines was reflected in the markedly different glycoproteomics signatures observed. Nevertheless, a restricted panel of SLeA-expressing glycoproteins with affinity for E-selectin remained conserved between the cell lines, consisting of a promising list of potentially targetable biomarkers for bladder cancer.

2020-05-11 | PXD016968 | Pride

Analysis of the substrate specificity of CpaA

Project description:Analysis of the substrates cleaveage specificity of the proteases CpaA from Acinetobacter.

2020-09-16 | PXD019941 | Pride

Urinary Glycosylated IGF2 peptides in association with CKD

Project description:Chronic kidney disease (CKD) is prevalent in 10% of world’s adult population, with Estimated Glomerular filtration rate (eGFR) and albuminuria employed in diagnosis. Role of protein glycosylation in causal mechanisms of CKD progression is largely unknown. Aim of this study was to identify urinary O-linked glycopeptides in association to CKD for better characterization of CKD molecular manifestations. Urine samples from 2 healthy and 8 CKD subjects were analyzed by CE-MS/MS and glycopeptide analysis using Proteome Discoverer 1.4. Distribution of identi-fied glycopeptides and correlation with Age, eGFR and Albuminuria were evaluated in 3810 da-tasets. In total, 17 O-linked glycopeptides from 7 different proteins were identified, derived primarily from Insulin-like growth factor-II (IGF2). Glycosylation occurred at the surface ex-posed IGF2 Threonine 96 position. Three glycopeptides (DVStPPTVLPDNFPRYPVGKF, DVStPPTVLPDNFPRYPVG and DVStPPTVLPDNFPRYP) exhibited positive correlation with Age. Glycopeptide (tPPTVLPDNFPRYP) showed strong negative association with eGFR. These results suggest that with aging and deteriorating kidney function, alterations in IGF2 proteoforms take place; which may reflect changes in mature IGF2 protein. Our results corroborate this hypothesis as IGF2 increased plasma levels were observed in CKD patients. Protease predictions, consider-ing also available transcriptomics data, suggest activation of cathepsin S with CKD, meriting further investigation.

2023-05-10 | PXD039829 | Pride

Mucin-type O-glycosylation Landscapes of SARS-CoV-2 Spike Proteins

Project description:The densely glycosylated spike (S) proteins that are highly exposed on the surface of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) facilitate viral attachment, entry, and membrane fusion. We have previously reported all the 22 N-glycosites and site-specific N-glycans in the S protein protomer. Herein, we report the comprehensive site-specific O-glycosylation landscapes of SARS-CoV-2 S proteins, which were characterized using high-resolution mass spectrometry. Following digestion using trypsin and trypsin/Glu-C, and de-N-glycosylation using PNGase F, we determined the mucin-type (GalNAc-type) O-glycosylation pattern of S proteins, including O-glycosites and the 6 most common O-glycans occupying them, via Byonic identification and manual validation. Finally, 43 O-glycosites were identified by higher energy collision-induced dissociation (HCD), and 11 O-glycosites were verified by electron transfer/higher energy collision-induced dissociation (EThcD) in the insect cell-expressed S protein. Most glycosites were modified by non-sialylated O-glycans such as HexNAc(1) and HexNAc(1)Hex(1). In contrast, 30 O-glycosites were identified by HCD, and 14 O-glycosites were verified by EThcD in the human cell-expressed S protein S1 subunit. Most glycosites were modified by sialylated O-glycans such as HexNAc(1)Hex(1)NeuAc(1) and HexNAc(1)Hex(1)NeuAc(2). Our results are the first to reveal that the SARS-CoV-2 S protein is a mucin-type glycoprotein; clustered O-glycans often occur in the N- and the C-termini of the S protein, and the O-glycosite and O-glycan compositions vary with the host cell type. These site-specific O-glycosylation landscapes of the SARS-CoV-2 S protein are expected to provide novel insights into the viral binding mechanism and present a strategy for the development of vaccines and targeted drugs.

2021-11-03 | PXD022896 | Pride

Glycoproteomics profiling of recombinant SARS-CoV-2 stabilized Spike construct

Project description:N-linked glycans of recombinant SARS-CoV-2 Spike protein with stabilizing mutations were profiled by LC-MS/MS.

2021-07-28 | PXD025554 | Pride

Installation of O-glycan sulfation capacities in human HEK293 cells

Project description:The human genome contains at least 35 genes that encode Golgi sulfotransferases functioning in the secretory pathway, where they are involved in decorating glycosaminoglycans, glycolipids, and glycoproteins with sulfate groups. Our insight into the sulfotransferases that serve glycoproteins and in particular GalNAc-type O-glycoproteins is limited, yet a number of important interactions with sulfated glycans by e.g. Selectins, Galectins, and Siglecs are thought to mainly rely on sulfated O-glycans. Moreover, sulfated mucins appear to be accumulated in respiratory diseases, arthritis and cancer. To explore further the genetic and biosynthetic regulation of sulfated O-glycans, we have started to expand a cell-based glycan array in the human HEK293 cell line with sulfation capacities. Here, we stably engineered O-glycan sulfation capacities in HEK293 cells by site-directed knock-in of sulfotransferase genes, in combination with knockout of endogenous core2 and/or sialylation capacities, to enable production of mucin reporters with sulfated O-glycans. Expression of GAL3ST2 in HEK293 cells resulted in sulfation of core1 and core2 O-glycans, whereas expression of GAL3ST4 resulted in sulfation of core1 only. We used the engineered cell library to dissect the binding specificity of galectin-4 and confirmed binding to the 3-O-sulfo-core1 O-glycan

2022-02-17 | PXD028982 | Pride

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data