This application proposes to study the biochemistry, molecular and cellular biology of type XVI collagen, a novel collagen discovered by cDNA cloning. the long term goal is to understand the biological role of this new collagen. Recently cDNA clones encoding the entire human alpha1(XVI) chain have been isolated and characterized. The overall structural arrangement of the deduced polypeptide differs significantly from other known collagen types. However, this polypeptide exhibits certain structural features characteristically seen in members of the fibril associated collagen, types IX, XII and XIV. It also resembles the cuticle collagen of C. elegans in that both collagens contain similar cysteine-rich motifs. In order to elucidate the biological function of this collagen, we propose the following specific aims. (1) Preparation and characterization of antibodies specific for the human alpha1(XVI) collagen using synthetic peptides derived from the cDNA sequence and recombinant proteins produced by expressing cDNA in cultured cells. (2) Characterization of type XVI collagen in cultured cells and tissues. The molecular size and chain composition of the native type XVI collagen will be determined by immunoblotting and immunoprecipitation using antibodies prepared above. (3) Elucidation of the spatial and temporal expression of the alpha1(XVI) collagen. The mRNA and protein expression in cells and tissues from humans and mice will be examined by RNA hybridization and immunofluorescence staining. (4) Elucidation of the structure/function relationship of the alpha1(VI) collagen using protein fragments, synthetic peptides and recombinant proteins produced in the mammalian expression system. (5) Characterization of other constituent chain(s) of type XVI collagen if studies proposed above indicate that it is a heterotrimer. (6) Isolation and characterization of the mouse alpha1(XVI) collagen gene and use the genomic fragment to develop transgenic mice with homozygous disruption of the alpha1(XVI) collagen gene.
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