Molecular assembly, secretion, and matrix deposition of type VI collagen.
Monoclonal antibodies reactive with the tissue form of type VI collagen were used to isolate the type VI collagen polypeptides from cultured fibroblasts and muscle cells. Two [35S]methionine-labeled polypeptides of 260 and 140 kD were found intracellularly, in the medium, and in the extracellular matrix of metabolically labeled cells. These polypeptides were disulfide cross-linked into very large complexes. The 260- and 140-kD polypeptides were intimately associated and could not be separated from each other by reduction without denaturation. In the absence of ascorbic acid, both polypeptides accumulated inside the cell, and their amounts in the medium and in the matrix were decreased. These results suggest that both the 260- and the 140-kD polypeptides are integral parts of the type VI collagen molecule. Examination of type VI collagen isolated from the intracellular pool by electron microscopy after rotary shadowing revealed structures corresponding to different stages of assembly of type VI collagen. Based on these images, a sequence for the intracellular assembly of type VI collagen could be discerned. Type VI collagen monomers are approximately 125 nm long and are composed of two globules separated by a thin strand. The monomers assemble into dimers and tetramers by lateral association. Only tetramers were present in culture media, whereas both tetramers and multimers were found in extracellular matrix extracts. The multimers appeared to have assembled from tetramers by end-to-end association into filaments that had prominent knobs and a periodicity of approximately 110 nm. These results show that, unlike other collagens, type VI collagen is assembled into tetramers before it is secreted from the cells, and they also suggest an extracellular aggregation mechanism that appears to be unique to this collagen.
Engvall
E
E
Hessle
H
H
Klier
G
G
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D001205:Ascorbic Acid / Q000494:pharmacology; D002478:Cells, Cultured; D003094:Collagen / Q000096:biosynthesis* / Q000276:immunology / Q000378:metabolism; D005109:Extracellular Matrix / Q000378:metabolism*; D005347:Fibroblasts / Q000187:drug effects / Q000378:metabolism / Q000648:ultrastructure; D006801:Humans; D007890:Leiomyosarcoma / Q000378:metabolism / Q000648:ultrastructure; D009132:Muscles / Q000187:drug effects / Q000378:metabolism / Q000648:ultrastructure; D012516:Osteosarcoma / Q000378:metabolism / Q000648:ultrastructure; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D051381:Rats; D012208:Rhabdomyosarcoma / Q000378:metabolism / Q000648:ultrastructure
D000911:Antibodies, Monoclonal; D003094:Collagen; D001205:Ascorbic Acid