title: A new method for detection of tumor driver-dependent changes of protein sialylation in a colon cancer cell line reveals nectin-3 as TGFBR2 target.
abstract: Protein-linked glycans play key roles in cell differentiation, cell-cell interactions, cell growth, adhesion and immune response. Aberrant glycosylation is a characteristic feature of tumor cells and is involved in tumor growth, escape from apoptosis, metastasis formation, and resistance to therapy. It can serve as cancer biomarker and treatment target. To enable comprehensive screening for the impact of tumor driving mutations in colorectal cancer cells we present a method for specific analysis of tumor driver-induced glycome changes. The strategy is based on a combination of three technologies, that is recombinase-mediated cassette exchange (RMCE), Click-It chemistry and mass spectrometry. The new method is exemplified by the analysis of the impact of inactivating mutations of the TGF-ß-receptor type II (TGFBR2) on sialic acid incorporation into protein-linked glycans of the colon cancer cell line HCT116. Overall, 70 proteins were found to show de novo sialic acid incorporation exclusively upon TGFBR2 expression whereas 7 proteins lost sialylation upon TGFBR2 reconstitution. Validation of detected candidate glycoproteins is demonstrated with the cell surface glycoprotein nectin-3 known to be involved in metastasis, invasion and prognosis of various cancers. Altogether, our new approach can help to systematically puzzle out the influence of tumor-specific mutations in a major signaling pathway, as exemplified by the TGFBR2 tumor suppressor, on the tumor glycome. It facilitates the identification of glycan-based tumor markers that could be used for diagnostic and therapeutic applications. In principle the outlined strategy can be adapted to any cancer cell line, tumor driver mutation and several glycan-building blocks.
authors.affiliation: Department of Applied Tumor Biology, Institute of Pathology, University Hospital Heidelberg, Heidelberg, Germany.|Department of Cancer Early Detection, German Cancer Research Center (DKFZ), Heidelberg, Germany.
authors.forename: Jennifer
authors.initials: J
authors.lastname: Lee
authors.affiliation: Department of Functional Proteome Analysis, German Cancer Research Center (DKFZ), Heidelberg, Germany.
authors.forename: Uwe
authors.initials: U
authors.lastname: Warnken
authors.affiliation: Department of Functional Proteome Analysis, German Cancer Research Center (DKFZ), Heidelberg, Germany.
authors.forename: Martina
authors.initials: M
authors.lastname: Schnölzer
authors.affiliation: Department of Applied Tumor Biology, Institute of Pathology, University Hospital Heidelberg, Heidelberg, Germany.|Department of Cancer Early Detection, German Cancer Research Center (DKFZ), Heidelberg, Germany.
authors.forename: Johannes
authors.initials: J
authors.lastname: Gebert
authors.affiliation: Department of Applied Tumor Biology, Institute of Pathology, University Hospital Heidelberg, Heidelberg, Germany.|Department of Cancer Early Detection, German Cancer Research Center (DKFZ), Heidelberg, Germany.
authors.forename: Jürgen
authors.initials: J
authors.lastname: Kopitz
mesh_terms: D001671:Biochemistry / Q000379:methods*; D015815:Cell Adhesion Molecules / Q000378:metabolism*; D045744:Cell Line, Tumor; D003110:Colonic Neoplasms / Q000503:physiopathology*; D006031:Glycosylation; D006801:Humans; D013058:Mass Spectrometry; D019158:N-Acetylneuraminic Acid / Q000737:chemistry*; D000075983:Nectins; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D000077294:Receptor, Transforming Growth Factor-beta Type II; D018125:Receptors, Transforming Growth Factor beta / Q000378:metabolism*
chemical_list: D015815:Cell Adhesion Molecules; C000619014:NECTIN3 protein, human; D000075983:Nectins; D018125:Receptors, Transforming Growth Factor beta; D017346:Protein Serine-Threonine Kinases; D000077294:Receptor, Transforming Growth Factor-beta Type II; D019158:N-Acetylneuraminic Acid
keywords: TGFBR2; colorectal cancer; microsatellite instability; nectin-3; sialylation