Transmembrane and secreted MUC1 probes show trafficking-dependent changes in O-glycan core profiles.
The human mucin MUC1 is expressed both as a transmembrane heterodimeric protein complex that recycles via the trans-Golgi network (TGN) and as a secreted isoform. To determine whether differences in cellular trafficking might influence the O-glycosylation profiles on these isoforms, we developed a model system consisting of membrane-bound and secretory-recombinant glycosylation probes. Secretory MUC1-S contains only a truncated repeat domain, whereas in MUC1-M constructs this domain is attached to the native transmembrane and cytoplasmic domains of MUC1 either directly (M0) or via an intermitting nonfunctional (M1) or functional sperm protein-enterokinase-agrin (SEA) module (M2); the SEA module contains a putative proteolytic cleavage site and is associated with proteins receiving extensive O-glycosylation. We showed that MUC1-M2 simulates endogenous MUC1 by recycling from the cell surface of Chinese hamster ovary (CHO) mutant ldlD14 cells through intracellular compartments where its glycosylation continues. The profiles of O-linked glycans on MUC1-S secreted by epithelial EBNA-293 and MCF-7 breast cancer cells revealed patterns dominated by core 2-based oligosaccharides. In contrast, the respective membrane-shed probes expressed in the same cells showed a complete shift to patterns dominated by sialyl core 1. In conclusion, glycan core profiles reflected the subcellular trafficking pathways of the secretory or membranous probes and the modifying activities of the resident glycosyltransferases.
Engelmann
Katja
K
Center of Biochemistry Medical Facility, University of Cologne, Joseph-Stelzmann-Str. 52, 50931 Köln, Germany.
Kinlough
Carol L
CL
Müller
Stefan
S
Razawi
Hani
H
Baldus
Stephan E
SE
Hughey
Rebecca P
RP
Hanisch
Franz-Georg
FG
D000595:Amino Acid Sequence; D000818:Animals; D001943:Breast Neoplasms / Q000378:metabolism*; D016466:CHO Cells; D045744:Cell Line, Tumor; D002462:Cell Membrane / Q000378:metabolism*; D002471:Cell Transformation, Neoplastic; D006224:Cricetinae; D005260:Female; D006031:Glycosylation; D006801:Humans; D008969:Molecular Sequence Data; D018396:Mucin-1 / Q000235:genetics / Q000378:metabolism*; D010446:Peptide Fragments / Q000235:genetics / Q000378:metabolism*; D011134:Polysaccharides / Q000737:chemistry / Q000378:metabolism*; D020033:Protein Isoforms; D021381:Protein Transport / Q000502:physiology; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012680:Sensitivity and Specificity; D016415:Sequence Alignment; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods; D013997:Time Factors; D021601:trans-Golgi Network / Q000378:metabolism
C116055:MUC1 tandem repeat peptide; D018396:Mucin-1; D010446:Peptide Fragments; D011134:Polysaccharides; D020033:Protein Isoforms; D011993:Recombinant Fusion Proteins