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Mucin biosynthesis and secretion in tracheal epithelial cells in primary culture

Svitacheva, N. LU and Davies, J. R. LU (2001) In Biochemical Journal 353(1). p.23-32
Abstract

Density-gradient centrifugation of bovine tracheal epithelial cell extracts revealed a 'high-density' (1.48 g/ml) sialic-acid-rich population as well as a 'low-density' (1.42 g/ml) one that reacted more strongly with a periodate-Schiff (PAS) assay. The sialic-acid-rich mucins were oligomeric molecules containing disulphide-bond-linked subunits and large glycosylated domains, whereas the PAS-reactive component seemed to be smaller and 'monomeric'. Only the 'high-density' population was secreted from cells cultured for 5 days on plastic or a collagen type 1, Matrigel or Vitrogen substrate. Release was less from cells grown on plastic than from those on a substrate and the amount was unaffected by increasing the thickness of the collagen... (More)

Density-gradient centrifugation of bovine tracheal epithelial cell extracts revealed a 'high-density' (1.48 g/ml) sialic-acid-rich population as well as a 'low-density' (1.42 g/ml) one that reacted more strongly with a periodate-Schiff (PAS) assay. The sialic-acid-rich mucins were oligomeric molecules containing disulphide-bond-linked subunits and large glycosylated domains, whereas the PAS-reactive component seemed to be smaller and 'monomeric'. Only the 'high-density' population was secreted from cells cultured for 5 days on plastic or a collagen type 1, Matrigel or Vitrogen substrate. Release was less from cells grown on plastic than from those on a substrate and the amount was unaffected by increasing the thickness of the collagen layer. For cells grown on collagen, the amount of the sialic-acid-rich mucin increased over 10 days, whereas the PAS-reactive component was largely absent after 24 h, which was consistent with an initial release of stored PAS-reactive molecules and synthesis of the sialic-acid-rich mucins de novo. Both [3H]proline and [35S]sulphate were poorly incorporated into mucins detected with the chemical assays but molecules with a higher buoyant density than that of either of the previously identified species were labelled with [35S]sulphate. The [35S]sulphate-labelled material yielded large trypsin-resistant fragments and contained O-linked glycans but was not affected by digestion with chondroitin ABC lyase or heparan sulphate lyase, suggesting that it is a mucin rather than a proteoglycan. [35S]Sulphate is thus a poor marker for the major oligomeric mucins produced by bovine tracheal epithelial cells but the radiolabel is incorporated into a heavily labelled mucin-like component.

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author
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organization
publishing date
type
Contribution to journal
publication status
published
subject
keywords
Airway, Bovine trachea, Cell culture, Mucus, Radio-labelling
in
Biochemical Journal
volume
353
issue
1
pages
23 - 32
publisher
Portland Press
external identifiers
  • pmid:11115395
  • scopus:0035152425
ISSN
0264-6021
DOI
10.1042/bj3530023
language
English
LU publication?
yes
id
4559df5f-8701-4830-bdca-17c346ecb511
date added to LUP
2020-02-25 16:29:52
date last changed
2021-02-17 05:54:03
@article{4559df5f-8701-4830-bdca-17c346ecb511,
  abstract     = {<p>Density-gradient centrifugation of bovine tracheal epithelial cell extracts revealed a 'high-density' (1.48 g/ml) sialic-acid-rich population as well as a 'low-density' (1.42 g/ml) one that reacted more strongly with a periodate-Schiff (PAS) assay. The sialic-acid-rich mucins were oligomeric molecules containing disulphide-bond-linked subunits and large glycosylated domains, whereas the PAS-reactive component seemed to be smaller and 'monomeric'. Only the 'high-density' population was secreted from cells cultured for 5 days on plastic or a collagen type 1, Matrigel or Vitrogen substrate. Release was less from cells grown on plastic than from those on a substrate and the amount was unaffected by increasing the thickness of the collagen layer. For cells grown on collagen, the amount of the sialic-acid-rich mucin increased over 10 days, whereas the PAS-reactive component was largely absent after 24 h, which was consistent with an initial release of stored PAS-reactive molecules and synthesis of the sialic-acid-rich mucins de novo. Both [<sup>3</sup>H]proline and [<sup>35</sup>S]sulphate were poorly incorporated into mucins detected with the chemical assays but molecules with a higher buoyant density than that of either of the previously identified species were labelled with [<sup>35</sup>S]sulphate. The [<sup>35</sup>S]sulphate-labelled material yielded large trypsin-resistant fragments and contained O-linked glycans but was not affected by digestion with chondroitin ABC lyase or heparan sulphate lyase, suggesting that it is a mucin rather than a proteoglycan. [<sup>35</sup>S]Sulphate is thus a poor marker for the major oligomeric mucins produced by bovine tracheal epithelial cells but the radiolabel is incorporated into a heavily labelled mucin-like component.</p>},
  author       = {Svitacheva, N. and Davies, J. R.},
  issn         = {0264-6021},
  language     = {eng},
  number       = {1},
  pages        = {23--32},
  publisher    = {Portland Press},
  series       = {Biochemical Journal},
  title        = {Mucin biosynthesis and secretion in tracheal epithelial cells in primary culture},
  url          = {http://dx.doi.org/10.1042/bj3530023},
  doi          = {10.1042/bj3530023},
  volume       = {353},
  year         = {2001},
}