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Formation of proteoglycan aggregates in rat chondrosarcoma chondrocyte cultures treated with tunicamycin

Lohmander, Stefan LU ; Fellini, S. A.; Kimura, J. H.; Stevens, R. L. and Hascall, V. C. (1983) In Journal of Biological Chemistry 258(20). p.12280-12286
Abstract

Proteoglycan monomer and link protein isolated from the Swarm rat chondrosarcoma both contain glycosylamine-linked oligosaccharides. In monomer, these N-linked oligosaccharides are concentrated in a region of the protein core which interacts specifically with both hyaluronate and link protein to form proteoglycan aggregates present in the cartilage matrix. Chondrocyte cultures were treated with tunicamycin to inhibit synthesis of the N-linked oligosaccharides, and the ability of the deficient proteoglycan and link protein to form aggregates was studied. Cultures were pretreated with tunicamycin for 3 h and then labeled with either [3H]mannose, [3H]glucosamine, [3H]serine, or with [35S]sulfate... (More)

Proteoglycan monomer and link protein isolated from the Swarm rat chondrosarcoma both contain glycosylamine-linked oligosaccharides. In monomer, these N-linked oligosaccharides are concentrated in a region of the protein core which interacts specifically with both hyaluronate and link protein to form proteoglycan aggregates present in the cartilage matrix. Chondrocyte cultures were treated with tunicamycin to inhibit synthesis of the N-linked oligosaccharides, and the ability of the deficient proteoglycan and link protein to form aggregates was studied. Cultures were pretreated with tunicamycin for 3 h and then labeled with either [3H]mannose, [3H]glucosamine, [3H]serine, or with [35S]sulfate for 6 h in the presence of tunicamycin. Formation of link protein-stabilized proteoglycan aggregates in the culture medium was inhibited by up to 40% when the cells were treated with 3 μg of tunicamycin/ml, a concentration which inhibited 3H incorporation with mannose as a precursor by about 90%, but by only 15% with glucosamine as a precursor. When exogenous proteoglycan aggregate was added to the culture medium, however, it was found that both endogenous monomer and link protein synthesized in the presence of tunicamycin were fully able to form link-stabilized aggregates. This suggests that glycosylamine-linked oligosaccharides on monomer and on link protein are not necessary for their specific interactions with hyaluronate and with each other. Further, although tunicamycin did not inhibit net synthesis of hyaluronate, transfer of hyaluronate from the cell layer to the culture medium was retarded. This phenomenon accounted for most if not all of the decrease in the amount of proteoglycan which formed aggregates in the medium of cultures treated with tunicamycin.

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published
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in
Journal of Biological Chemistry
volume
258
issue
20
pages
7 pages
publisher
American Society for Biochemistry and Molecular Biology
external identifiers
  • Scopus:0021052508
ISSN
0021-9258
language
English
LU publication?
yes
id
b5a07ed1-cf82-4b97-8c49-d5c79fe1cfc5
date added to LUP
2016-05-04 12:55:37
date last changed
2016-07-04 11:31:14
@misc{b5a07ed1-cf82-4b97-8c49-d5c79fe1cfc5,
  abstract     = {<p>Proteoglycan monomer and link protein isolated from the Swarm rat chondrosarcoma both contain glycosylamine-linked oligosaccharides. In monomer, these N-linked oligosaccharides are concentrated in a region of the protein core which interacts specifically with both hyaluronate and link protein to form proteoglycan aggregates present in the cartilage matrix. Chondrocyte cultures were treated with tunicamycin to inhibit synthesis of the N-linked oligosaccharides, and the ability of the deficient proteoglycan and link protein to form aggregates was studied. Cultures were pretreated with tunicamycin for 3 h and then labeled with either [<sup>3</sup>H]mannose, [<sup>3</sup>H]glucosamine, [<sup>3</sup>H]serine, or with [<sup>35</sup>S]sulfate for 6 h in the presence of tunicamycin. Formation of link protein-stabilized proteoglycan aggregates in the culture medium was inhibited by up to 40% when the cells were treated with 3 μg of tunicamycin/ml, a concentration which inhibited <sup>3</sup>H incorporation with mannose as a precursor by about 90%, but by only 15% with glucosamine as a precursor. When exogenous proteoglycan aggregate was added to the culture medium, however, it was found that both endogenous monomer and link protein synthesized in the presence of tunicamycin were fully able to form link-stabilized aggregates. This suggests that glycosylamine-linked oligosaccharides on monomer and on link protein are not necessary for their specific interactions with hyaluronate and with each other. Further, although tunicamycin did not inhibit net synthesis of hyaluronate, transfer of hyaluronate from the cell layer to the culture medium was retarded. This phenomenon accounted for most if not all of the decrease in the amount of proteoglycan which formed aggregates in the medium of cultures treated with tunicamycin.</p>},
  author       = {Lohmander, Stefan and Fellini, S. A. and Kimura, J. H. and Stevens, R. L. and Hascall, V. C.},
  issn         = {0021-9258},
  language     = {eng},
  number       = {20},
  pages        = {12280--12286},
  publisher    = {ARRAY(0x94b5e58)},
  series       = {Journal of Biological Chemistry},
  title        = {Formation of proteoglycan aggregates in rat chondrosarcoma chondrocyte cultures treated with tunicamycin},
  volume       = {258},
  year         = {1983},
}