Recombinant MUC1 probe authentically reflects cell-specific O-glycosylation profiles of endogenous breast cancer mucin. High density and prevalent core 2-based glycosylation.

Abstract

Knowledge about the O-linked glycan chains of tumor-associated MUC1 is primarily based on enzymatic and immunochemical evidence. To obtain structural information and to overcome limitations by the scarcity of endogenous mucin, we expressed a recombinant glycosylation probe corresponding to six MUC1 tandem repeats in four breast cancer cell lines. Comparative analyses of the O-glycan profiles were performed after hydrazinolysis and normal phase chromatography of 2-aminobenzamide-labeled glycans. Except for a general reduction in the O-glycan chain lengths and a high density glycosylation, no common structural pattern was revealed. T47D fusion protein exhibits an almost complete shift from core 2 to core 1 expression with a preponderance of sialylated glycans. By contrast, MCF-7, MDA-MB231, and ZR75-1 cells glycosylate the MUC1 repeat peptide preferentially with core 2-based glycans terminating mostly with alpha 3-linked sialic acid (MDA-MB231, ZR75-1) or alpha 2/3-linked fucose (MCF-7). Endogenous MUC1 from T47D and MCF-7 cell supernatants revealed almost identical O-glycosylation profiles compared with the respective recombinant probes, indicating that the fusion proteins reflected the authentic O-glycan profiles of the cells. The structural patterns in the majority of cells under study are in conflict with biosynthetic models of MUC1 O-glycosylation in breast cancer, which claim that the truncation of normal core 2-based polylactosamine structures to short sialylated core 1-based glycans is due to the reduced activity of core 2-forming beta 6-N-acetylglucosaminyltransferases and/or to overexpression of competitive alpha 3- sialyltransferase.

• Authors: Stefan Müller, Franz-Georg Hanisch
• Journal: The Journal of biological chemistry (J Biol Chem) Vol. 277 Issue 29 Pg. 26103-12 (Jul 19 2002) ISSN: 0021-9258 [Print] United States
• PMID: 12000758 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't, Research Support, U.S. Gov't, P.H.S.)

Chemical References

• MUC1 tandem repeat peptide
• Mucin-1
• Peptide Fragments

Topics

• Amino Acid Sequence
• Breast Neoplasms (metabolism)
• Carbohydrate Conformation
• Chromatography, High Pressure Liquid
• Female
• Glycosylation
• Humans
• Mass Spectrometry
• Molecular Sequence Data
• Mucin-1 (genetics, metabolism)
• Peptide Fragments (genetics, metabolism)
• Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
• Tandem Repeat Sequences
• Tumor Cells, Cultured