N-Glycosylation affects the stability and barrier function of the MUC16 mucin

J Biol Chem. 2017 Jun 30;292(26):11079-11090. doi: 10.1074/jbc.M116.770123. Epub 2017 May 9.

Abstract

Transmembrane mucins are highly O-glycosylated glycoproteins that coat the apical glycocalyx on mucosal surfaces and represent the first line of cellular defense against infection and injury. Relatively low levels of N-glycans are found on transmembrane mucins, and their structure and function remain poorly characterized. We previously reported that carbohydrate-dependent interactions of transmembrane mucins with galectin-3 contribute to maintenance of the epithelial barrier at the ocular surface. Now, using MALDI-TOF mass spectrometry, we report that transmembrane mucin N-glycans in differentiated human corneal epithelial cells contain primarily complex-type structures with N-acetyllactosamine, a preferred galectin ligand. In N-glycosylation inhibition experiments, we find that treatment with tunicamycin and siRNA-mediated knockdown of the Golgi N-acetylglucosaminyltransferase I gene (MGAT1) induce partial loss of both total and cell-surface levels of the largest mucin, MUC16, and a concomitant reduction in glycocalyx barrier function. Moreover, we identified a distinct role for N-glycans in promoting MUC16's binding affinity toward galectin-3 and in causing retention of the lectin on the epithelial cell surface. Taken together, these studies define a role for N-linked oligosaccharides in supporting the stability and function of transmembrane mucins on mucosal surfaces.

Keywords: N-linked glycosylation; epithelial cell; galectin; glycosyltransferase; mucin.

MeSH terms

  • Blood Proteins
  • CA-125 Antigen / genetics
  • CA-125 Antigen / metabolism*
  • Cell Line, Transformed
  • Cornea / metabolism*
  • Epithelial Cells / metabolism*
  • Galectin 3 / genetics
  • Galectin 3 / metabolism*
  • Galectins
  • Glycocalyx / genetics
  • Glycocalyx / metabolism*
  • Glycosylation
  • Humans
  • Membrane Proteins / genetics
  • Membrane Proteins / metabolism*
  • Protein Stability

Substances

  • Blood Proteins
  • CA-125 Antigen
  • Galectin 3
  • Galectins
  • LGALS3 protein, human
  • MUC16 protein, human
  • Membrane Proteins