Stable expression of chicken type-VI collagen alpha 1, alpha 2 and alpha 3 cDNAs in murine NIH/3T3 cells.
As a component of an extensive network of microfibrils interwoven with large collagen fibers and in close contact with cell surfaces, type VI collagen plays an important role in cell-matrix interactions. To investigate the behaviour of chicken type VI collagen chains in heterologous host cells as a means to understanding the pattern of assembly of this collagen, we transfected murine NIH/3T3 cells with cDNAs encoding chicken alpha 1(VI), alpha 2(VI) and alpha 3(VI) chains. Cell lines that constitutively expressed the individual chains were analyzed by metabolic labeling and immunoprecipitation with specific antibodies. No self-association was observed for either alpha 1(VI) or alpha 2(VI) chains which were secreted as monomeric polypeptides. Furthermore, neither the chicken alpha 1(VI) nor alpha 2(VI) chains associated with the endogenous murine chains to form chimeric chicken/murine heterotrimers. In contrast, chimeric chicken/murine heterotrimers were detected in cell lines transfected with chicken alpha 3(VI) cDNA. These chimeric forms appeared to be properly aligned since their triple helices were stable to pepsin digestion. In addition, the chimeric heterotrimers coassembled and gave rise to disulfide-linked type VI collagen molecules.
Colombatti
A
A
Divisione di Oncologia Sperimentale 2, Centro di Riferimento Oncologico, Aviano, Italy.
Bonaldo
P
P
Bucciotti
F
F
D016475:3T3 Cells; D000818:Animals; D015152:Blotting, Northern; D002645:Chickens; D003094:Collagen / Q000096:biosynthesis* / Q000235:genetics* / Q000302:isolation & purification; D004247:DNA / Q000235:genetics*; D004591:Electrophoresis, Polyacrylamide Gel; D005455:Fluorescent Antibody Technique; D046911:Macromolecular Substances; D008715:Methionine / Q000378:metabolism; D051379:Mice; D012313:RNA / Q000235:genetics / Q000302:isolation & purification; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000302:isolation & purification; D014162:Transfection
D046911:Macromolecular Substances; D011994:Recombinant Proteins; D012313:RNA; D003094:Collagen; D004247:DNA; D008715:Methionine