O-Linked-N-acetylglucosamine on extracellular protein domains mediates epithelial cell–matrix interactions

Sakaidani, Yuta; Nomura, Tomoko; Matsuura, Aiko; Ito, Makiko; Suzuki, Emiko; Murakami, Kosuke; Nadano, Daita; Matsuda, Tsukasa; Furukawa, Koichi; Okajima, Tetsuya

doi:10.1038/ncomms1591

Cited by 163 publications

(185 citation statements)

References 40 publications

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“…Among the proteins in both the human and mouse proteome that contain this motif, ∼30% are localized within the ECM. The importance of this modification within the ECM has been demonstrated in Drosophila, where loss of EOGT causes defects in the apical ECM (37). The proteins containing the CXXGXS/TGXXC motif appear conserved across species, with 83 proteins conserved between human and mouse, and 15 of 18 Drosophila proteins having orthologs in both human and mouse.…”

Section: O-glcnacylation On a Secreted Protein And On The Extracellularmentioning

confidence: 99%

“…from Drosophila to mammals (37,38). There is no evidence that intracellular forms of OGT or O-GlcNAcase occur in the extracellular or luminal spaces.…”

Section: O-glcnacylation On a Secreted Protein And On The Extracellularmentioning

confidence: 99%

“…There is no evidence that intracellular forms of OGT or O-GlcNAcase occur in the extracellular or luminal spaces. Four O-GlcNAcylated peptides from these proteins were located within an EGF-like domain, sharing a similar motif sequence CXXGXS/TGXXC to the reported extracellular O-GlcNAcylation on NOTCH and Dumpy in Drosophila and Notch1 in mammals (36)(37)(38). We also identified an O-GlcNAc site on the EGF-like domain of NOTCH2 protein of the Notch signaling pathway, which we infer is on the T residue in the YSCVCSPGFTGQR sequence, consistent with the CXXGXS/ TGXXC motif (Table 3).…”

Section: O-glcnacylation On a Secreted Protein And On The Extracellularmentioning

confidence: 99%

“…It resides in the endoplasmic reticulum (ER) within the secretory pathway and is termed EGF domain-specific O-GlcNAc transferase (EOGT) (36,37). EOGT is conserved Proteins are shaded in different colors according to their functional category.…”

Section: O-glcnacylation On a Secreted Protein And On The Extracellularmentioning

confidence: 99%

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Tandem mass spectrometry identifies many mouse brain O -GlcNAcylated proteins including EGF domain-specific O -GlcNAc transferase targets

Alfaro

Gong

Monroe

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

260

265

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O -linked N -acetylglucosamine ( O -GlcNAc) is a reversible posttranslational modification of Ser and Thr residues on cytosolic and nuclear proteins of higher eukaryotes catalyzed by O -GlcNAc transferase (OGT). O -GlcNAc has recently been found on Notch1 extracellular domain catalyzed by EGF domain-specific OGT. Aberrant O -GlcNAc modification of brain proteins has been linked to Alzheimer's disease (AD). However, understanding specific functions of O -GlcNAcylation in AD has been impeded by the difficulty in characterization of O -GlcNAc sites on proteins. In this study, we modified a chemical/enzymatic photochemical cleavage approach for enriching O -GlcNAcylated peptides in samples containing ∼100 μg of tryptic peptides from mouse cerebrocortical brain tissue. A total of 274 O -GlcNAcylated proteins were identified. Of these, 168 were not previously known to be modified by O -GlcNAc. Overall, 458 O -GlcNAc sites in 195 proteins were identified. Many of the modified residues are either known phosphorylation sites or located proximal to known phosphorylation sites. These findings support the proposed regulatory cross-talk between O -GlcNAcylation and phosphorylation. This study produced the most comprehensive O -GlcNAc proteome of mammalian brain tissue with both protein identification and O -GlcNAc site assignment. Interestingly, we observed O -β-GlcNAc on EGF-like repeats in the extracellular domains of five membrane proteins, expanding the evidence for extracellular O -GlcNAcylation by the EGF domain-specific OGT. We also report a GlcNAc-β-1,3-Fuc-α-1- O -Thr modification on the EGF-like repeat of the versican core protein, a proposed substrate of Fringe β-1,3- N -acetylglucosaminyltransferases.

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Section: O-glcnacylation On a Secreted Protein And On The Extracellularmentioning

confidence: 99%

“…from Drosophila to mammals (37,38). There is no evidence that intracellular forms of OGT or O-GlcNAcase occur in the extracellular or luminal spaces.…”

Section: O-glcnacylation On a Secreted Protein And On The Extracellularmentioning

confidence: 99%

Section: O-glcnacylation On a Secreted Protein And On The Extracellularmentioning

confidence: 99%

Section: O-glcnacylation On a Secreted Protein And On The Extracellularmentioning

confidence: 99%

See 2 more Smart Citations

Tandem mass spectrometry identifies many mouse brain O -GlcNAcylated proteins including EGF domain-specific O -GlcNAc transferase targets

Alfaro

Gong

Monroe

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

260

265

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“…The third type of O-glycosylation that occurs on EGF repeats is O-GlcNAcylation, which is mediated by the EGF domainspecific O-GlcNAc transferase (Eogt in flies, EOGT1 in mammals) (36,37). EOGT is localized to the endoplasmic reticulum and modifies secreted and membrane proteins, making it distinct from the nuclear and cytoplasmic O-GlcNAc transferase (36).…”

mentioning

confidence: 99%

Mapping Sites of O-Glycosylation and Fringe Elongation on Drosophila Notch

Harvey

Rana

Moss

et al. 2016

Journal of Biological Chemistry

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Glycosylation of the Notch receptor is essential for its activity and serves as an important modulator of signaling. Three major forms of O-glycosylation are predicted to occur at consensus sites within the epidermal growth factor-like repeats in the extracellular domain of the receptor: O-fucosylation, O-glucosylation, and O-GlcNAcylation. We have performed comprehensive mass spectral analyses of these three types of O-glycosylation on Drosophila Notch produced in S2 cells and identified peptides containing all 22 predicted O-fucose sites, all 18 predicted O-glucose sites, and all 18 putative O-GlcNAc sites. Using semiquantitative mass spectral methods, we have evaluated the occupancy and relative amounts of glycans at each site. The majority of the O-fucose sites were modified to high stoichiometries. Upon expression of the ␤3-N-acetylglucosaminyltransferase Fringe with Notch, we observed varying degrees of elongation beyond O-fucose monosaccharide, indicating that Fringe preferentially modifies certain sites more than others. Rumi modified O-glucose sites to high stoichiometries, although elongation of the O-glucose was site-specific. Although the current putative consensus sequence for O-GlcNAcylation predicts 18 O-GlcNAc sites on Notch, we only observed apparent O-GlcNAc modification at five sites. In addition, we performed mass spectral analysis on endogenous Notch purified from Drosophila embryos and found that the glycosylation states were similar to those found on Notch from S2 cells. These data provide foundational information for future studies investigating the mechanisms of how O-glycosylation regulates Notch activity.

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Multiple roles for O‐glycans in Notch signalling

Varshney

Stanley

2018

FEBS Letters

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Notch signalling regulates a plethora of developmental processes and is also essential for the maintenance of tissue homeostasis in adults. Therefore, fine-tuning of Notch signalling strength needs to be tightly regulated. Of key importance for the regulation of Notch signalling are O-fucose, O-GlcNAc and O-glucose glycans attached to the extracellular domain of Notch receptors. The EGF repeats of the Notch receptor extracellular domain harbour consensus sites for addition of the different types of O-glycan to Ser or Thr, which takes place in the endoplasmic reticulum. Studies from Drosophila to mammals have demonstrated the multifaceted roles of O-glycosylation in regulating Notch signalling. O-glycosylation modulates different aspects of Notch signalling including recognition by Notch ligands, the strength of ligand binding, Notch receptor trafficking, stability and activation at the cell surface. Defects in O-glycosylation of Notch receptors give rise to pathologies in humans. This Review summarizes the nature of the O-glycans on Notch receptors and their differential effects on Notch signalling.

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O-Linked-N-acetylglucosamine on extracellular protein domains mediates epithelial cell–matrix interactions

Cited by 163 publications

References 40 publications

Tandem mass spectrometry identifies many mouse brain O -GlcNAcylated proteins including EGF domain-specific O -GlcNAc transferase targets

Tandem mass spectrometry identifies many mouse brain O -GlcNAcylated proteins including EGF domain-specific O -GlcNAc transferase targets

Mapping Sites of O-Glycosylation and Fringe Elongation on Drosophila Notch

Multiple roles for O‐glycans in Notch signalling

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