title: N-glycosylation Profiling of Colorectal Cancer Cell Lines Reveals Association of Fucosylation with Differentiation and Caudal Type Homebox 1 (CDX1)/Villin mRNA Expression.
abstract: Various cancers such as colorectal cancer (CRC) are associated with alterations in protein glycosylation. CRC cell lines are frequently used to study these (glyco)biological changes and their mechanisms. However, differences between CRC cell lines with regard to their glycosylation have hitherto been largely neglected. Here, we comprehensively characterized the N-glycan profiles of 25 different CRC cell lines, derived from primary tumors and metastatic sites, in order to investigate their potential as glycobiological tumor model systems and to reveal glycans associated with cell line phenotypes. We applied an optimized, high-throughput membrane-based enzymatic glycan release for small sample amounts. Released glycans were derivatized to stabilize and differentiate between α2,3- and α2,6-linked N-acetylneuraminic acids, followed by N-glycosylation analysis by MALDI-TOF(/TOF)-MS. Our results showed pronounced differences between the N-glycosylation patterns of CRC cell lines. CRC cell line profiles differed from tissue-derived N-glycan profiles with regard to their high-mannose N-glycan content but showed a large overlap for complex type N-glycans, supporting their use as a glycobiological cancer model system. Importantly, we could show that the high-mannose N-glycans did not only occur as intracellular precursors but were also present at the cell surface. The obtained CRC cell line N-glycan features were not clearly correlated with mRNA expression levels of glycosyltransferases, demonstrating the usefulness of performing the structural analysis of glycans. Finally, correlation of CRC cell line glycosylation features with cancer cell markers and phenotypes revealed an association between highly fucosylated glycans and CDX1 and/or villin mRNA expression that both correlate with cell differentiation. Together, our findings provide new insights into CRC-associated glycan changes and setting the basis for more in-depth experiments on glycan function and regulation.
authors.affiliation: From the ‡Center for Proteomics and Metabolomics.
authors.forename: Stephanie
authors.initials: S
authors.lastname: Holst
authors.affiliation: From the ‡Center for Proteomics and Metabolomics.
authors.forename: Anna J M
authors.initials: AJ
authors.lastname: Deuss
authors.affiliation: §Department of Surgery, and.
authors.forename: Gabi W
authors.initials: GW
authors.lastname: van Pelt
authors.affiliation: ‖Department of Molecular Cell Biology and Immunology and.
authors.forename: Sandra J
authors.initials: SJ
authors.lastname: van Vliet
authors.affiliation: ‖Department of Molecular Cell Biology and Immunology and.
authors.forename: Juan J
authors.initials: JJ
authors.lastname: Garcia-Vallejo
authors.affiliation: From the ‡Center for Proteomics and Metabolomics.
authors.forename: Carolien A M
authors.initials: CA
authors.lastname: Koeleman
authors.affiliation: From the ‡Center for Proteomics and Metabolomics.
authors.forename: André M
authors.initials: AM
authors.lastname: Deelder
authors.affiliation: §Department of Surgery, and.
authors.forename: Wilma E
authors.initials: WE
authors.lastname: Mesker
authors.affiliation: §Department of Surgery, and.
authors.forename: Rob A
authors.initials: RA
authors.lastname: Tollenaar
authors.affiliation: From the ‡Center for Proteomics and Metabolomics, ¶Department of RheumatologyLeiden University Medical Center, Leiden, The Netherlands; ‡‡Univ. Lille, CNRS, UMR 8576, UGSF, Unité de Glycobiologie Structurale et Fonctionnelle, F 59 000 Lille, France.
authors.forename: Yoann
authors.initials: Y
authors.lastname: Rombouts
authors.affiliation: From the ‡Center for Proteomics and Metabolomics, ‖Department of Molecular Cell Biology and Immunology and **Division of BioAnalytical Chemistry, VU University Medical Center, Amsterdam, The Netherlands; m.wuhrer@lumc.nl.
authors.forename: Manfred
authors.initials: M
authors.lastname: Wuhrer
mesh_terms: D018938:Caco-2 Cells; D002454:Cell Differentiation / Q000235:genetics; D045744:Cell Line, Tumor; D015179:Colorectal Neoplasms / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D005643:Fucose / Q000378:metabolism; D015972:Gene Expression Regulation, Neoplastic; D054794:Glycomics / Q000379:methods*; D006031:Glycosylation; D016695:Glycosyltransferases / Q000235:genetics / Q000378:metabolism; D045325:HCT116 Cells; D019073:HT29 Cells; D018398:Homeodomain Proteins / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008840:Microfilament Proteins / Q000235:genetics / Q000378:metabolism*; D011134:Polysaccharides / Q000378:metabolism*; D025341:Principal Component Analysis; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D053719:Tandem Mass Spectrometry
chemical_list: C491273:CDX1 protein, human; D018398:Homeodomain Proteins; D008840:Microfilament Proteins; D011134:Polysaccharides; D012333:RNA, Messenger; C026615:villin; D005643:Fucose; D016695:Glycosyltransferases