Precision mapping of the human O-GalNAc glycoproteome through SimpleCell technology

Catharina Steentoft; Sergey Y Vakhrushev; Hiren J Joshi; Yun Kong; Malene B Vester-Christensen; Katrine Ter-Borch Gram Schjoldager; Kirstine Lavrsen; Sally Dabelsteen; Nis Borbye Pedersen; Lara Patricia Marcos da Silva; Ramneek Gupta; Eric Paul Bennett; Ulla Mandel; Søren Brunak; Hans H Wandall; Steven B Levery; Henrik Clausen

doi:10.1038/emboj.2013.79

Precision mapping of the human O-GalNAc glycoproteome through SimpleCell technology

Catharina Steentoft, Sergey Y Vakhrushev, Hiren J Joshi, Yun Kong, Malene B Vester-Christensen, Katrine Ter-Borch Gram Schjoldager, Kirstine Lavrsen, Sally Dabelsteen, Nis Borbye Pedersen, Lara Patricia Marcos da Silva, Ramneek Gupta, Eric Paul Bennett, Ulla Mandel, Søren Brunak, Hans H Wandall, Steven B Levery, Henrik Clausen

709 Citations (Scopus)

Abstract

Glycosylation is the most abundant and diverse posttranslational modification of proteins. While several types of glycosylation can be predicted by the protein sequence context, and substantial knowledge of these glycoproteomes is available, our knowledge of the GalNAc-type O-glycosylation is highly limited. This type of glycosylation is unique in being regulated by 20 polypeptide GalNAc-transferases attaching the initiating GalNAc monosaccharides to Ser and Thr (and likely some Tyr) residues. We have developed a genetic engineering approach using human cell lines to simplify O-glycosylation (SimpleCells) that enables proteome-wide discovery of O-glycan sites using 'bottom-up' ETD-based mass spectrometric analysis. We implemented this on 12 human cell lines from different organs, and present a first map of the human O-glycoproteome with almost 3000 glycosites in over 600 O-glycoproteins as well as an improved NetOGlyc4.0 model for prediction of O-glycosylation. The finding of unique subsets of O-glycoproteins in each cell line provides evidence that the O-glycoproteome is differentially regulated and dynamic. The greatly expanded view of the O-glycoproteome should facilitate the exploration of how site-specific O-glycosylation regulates protein function.

Original language	English
Journal	E M B O Journal
Volume	32
Issue number	10
Pages (from-to)	1478-1488
Number of pages	11
ISSN	0261-4189
DOIs	https://doi.org/10.1038/emboj.2013.79
Publication status	Published - 15 May 2013

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Steentoft, C., Vakhrushev, S. Y., Joshi, H. J., Kong, Y., Vester-Christensen, M. B., Schjoldager, K. T-B. G., Lavrsen, K., Dabelsteen, S., Pedersen, N. B., da Silva, L. P. M., Gupta, R., Paul Bennett, E., Mandel, U., Brunak, S., Wandall, H. H., Levery, S. B., & Clausen, H. (2013). Precision mapping of the human O-GalNAc glycoproteome through SimpleCell technology. E M B O Journal, 32(10), 1478-1488. https://doi.org/10.1038/emboj.2013.79

Steentoft, C, Vakhrushev, SY , Joshi, HJ, Kong, Y, Vester-Christensen, MB, Schjoldager, KT-BG, Lavrsen, K, Dabelsteen, S, Pedersen, NB, da Silva, LPM, Gupta, R, Paul Bennett, E , Mandel, U , Brunak, S , Wandall, HH, Levery, SB & Clausen, H 2013, 'Precision mapping of the human O-GalNAc glycoproteome through SimpleCell technology', E M B O Journal, vol. 32, no. 10, pp. 1478-1488. https://doi.org/10.1038/emboj.2013.79

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title = "Precision mapping of the human O-GalNAc glycoproteome through SimpleCell technology",

abstract = "Glycosylation is the most abundant and diverse posttranslational modification of proteins. While several types of glycosylation can be predicted by the protein sequence context, and substantial knowledge of these glycoproteomes is available, our knowledge of the GalNAc-type O-glycosylation is highly limited. This type of glycosylation is unique in being regulated by 20 polypeptide GalNAc-transferases attaching the initiating GalNAc monosaccharides to Ser and Thr (and likely some Tyr) residues. We have developed a genetic engineering approach using human cell lines to simplify O-glycosylation (SimpleCells) that enables proteome-wide discovery of O-glycan sites using 'bottom-up' ETD-based mass spectrometric analysis. We implemented this on 12 human cell lines from different organs, and present a first map of the human O-glycoproteome with almost 3000 glycosites in over 600 O-glycoproteins as well as an improved NetOGlyc4.0 model for prediction of O-glycosylation. The finding of unique subsets of O-glycoproteins in each cell line provides evidence that the O-glycoproteome is differentially regulated and dynamic. The greatly expanded view of the O-glycoproteome should facilitate the exploration of how site-specific O-glycosylation regulates protein function.",

author = "Catharina Steentoft and Vakhrushev, {Sergey Y} and Joshi, {Hiren J} and Yun Kong and Vester-Christensen, {Malene B} and Schjoldager, {Katrine Ter-Borch Gram} and Kirstine Lavrsen and Sally Dabelsteen and Pedersen, {Nis Borbye} and {da Silva}, {Lara Patricia Marcos} and Ramneek Gupta and {Paul Bennett}, Eric and Ulla Mandel and S{\o}ren Brunak and Wandall, {Hans H} and Levery, {Steven B} and Henrik Clausen",

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AU - Steentoft, Catharina

AU - Vakhrushev, Sergey Y

AU - Joshi, Hiren J

AU - Kong, Yun

AU - Vester-Christensen, Malene B

AU - Schjoldager, Katrine Ter-Borch Gram

AU - Lavrsen, Kirstine

AU - Dabelsteen, Sally

AU - Pedersen, Nis Borbye

AU - da Silva, Lara Patricia Marcos

AU - Gupta, Ramneek

AU - Paul Bennett, Eric

AU - Mandel, Ulla

AU - Brunak, Søren

AU - Wandall, Hans H

AU - Levery, Steven B

AU - Clausen, Henrik

PY - 2013/5/15

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N2 - Glycosylation is the most abundant and diverse posttranslational modification of proteins. While several types of glycosylation can be predicted by the protein sequence context, and substantial knowledge of these glycoproteomes is available, our knowledge of the GalNAc-type O-glycosylation is highly limited. This type of glycosylation is unique in being regulated by 20 polypeptide GalNAc-transferases attaching the initiating GalNAc monosaccharides to Ser and Thr (and likely some Tyr) residues. We have developed a genetic engineering approach using human cell lines to simplify O-glycosylation (SimpleCells) that enables proteome-wide discovery of O-glycan sites using 'bottom-up' ETD-based mass spectrometric analysis. We implemented this on 12 human cell lines from different organs, and present a first map of the human O-glycoproteome with almost 3000 glycosites in over 600 O-glycoproteins as well as an improved NetOGlyc4.0 model for prediction of O-glycosylation. The finding of unique subsets of O-glycoproteins in each cell line provides evidence that the O-glycoproteome is differentially regulated and dynamic. The greatly expanded view of the O-glycoproteome should facilitate the exploration of how site-specific O-glycosylation regulates protein function.

AB - Glycosylation is the most abundant and diverse posttranslational modification of proteins. While several types of glycosylation can be predicted by the protein sequence context, and substantial knowledge of these glycoproteomes is available, our knowledge of the GalNAc-type O-glycosylation is highly limited. This type of glycosylation is unique in being regulated by 20 polypeptide GalNAc-transferases attaching the initiating GalNAc monosaccharides to Ser and Thr (and likely some Tyr) residues. We have developed a genetic engineering approach using human cell lines to simplify O-glycosylation (SimpleCells) that enables proteome-wide discovery of O-glycan sites using 'bottom-up' ETD-based mass spectrometric analysis. We implemented this on 12 human cell lines from different organs, and present a first map of the human O-glycoproteome with almost 3000 glycosites in over 600 O-glycoproteins as well as an improved NetOGlyc4.0 model for prediction of O-glycosylation. The finding of unique subsets of O-glycoproteins in each cell line provides evidence that the O-glycoproteome is differentially regulated and dynamic. The greatly expanded view of the O-glycoproteome should facilitate the exploration of how site-specific O-glycosylation regulates protein function.

U2 - 10.1038/emboj.2013.79

DO - 10.1038/emboj.2013.79

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EP - 1488

JO - E M B O Journal

JF - E M B O Journal

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ER -

Precision mapping of the human O-GalNAc glycoproteome through SimpleCell technology

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