Secretion and matrix assembly of recombinant type VI collagen.
A monomer of type VI collagen is composed of three different chains of 140 (alpha 1), 130 (alpha 2), and 250-350 kDa (alpha 3). Monomers assemble into dimers (6 chains) and tetramers (12 chains) that are stabilized by disulfide bonds and, once associated one to another, give rise to a microfilamentous network in close apposition with cell surfaces and banded collagen fibers. We have derived murine NIH/3T3 cell lines that were transfected with the cDNAs for the three chains and that constitutively expressed chicken type VI collagen. Cotransfection was efficient because, in three out of six isolated cell lines, all chicken chains were expressed. Southern blotting demonstrated that several copies of each cDNA were integrated approximately in equal number. Expression of the three polypeptide chains was consistent with the levels of the respective mRNAs. The three chicken chains assembled by disulfide bonding to form correctly folded triple helical aggregated composites with sizes corresponding to type VI collagen monomers, dimers, and tetramers. These functional recombinant assemblies were secreted and became incorporated into the extracellular matrix, where they formed an extensive fibrillar network.
Colombatti
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A
Divisione di Oncologia Sperimentale 2, Centro di Riferimento Oncologico, Aviano, Italy.
Mucignat
M T
MT
Bonaldo
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D016475:3T3 Cells; D000818:Animals; D015152:Blotting, Northern; D002645:Chickens; D003001:Cloning, Molecular; D003094:Collagen / Q000235:genetics / Q000378:metabolism*; D018076:DNA, Complementary; D005109:Extracellular Matrix / Q000378:metabolism*; D051379:Mice; D011233:Precipitin Tests; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D014162:Transfection
D018076:DNA, Complementary; D012333:RNA, Messenger; D011994:Recombinant Proteins; D003094:Collagen