Differential expression of two basement membrane collagen genes, COL4A6 and COL4A5, demonstrated by immunofluorescence staining using peptide-specific monoclonal antibodies

Genes for the human α5(IV) and α6(IV) collagen chains have a unique arrangement in that they are colocalized on chromosome Xq22 in a head-to-head fashion and appear to share a common bidirectional promoter. In addition we reported a novel observation that the COL4A6 gene is transcribed from two alternative promoters in a tissue-specific manner (Sugimoto, M., T. Oohashi, and Y. Ninomiya. 1994. Proc. Natl. Acad. Sci. USA. 91:11679-11683). To know whether the translation products of both genes are colocalized in various tissues, we raised α5(IV) and α6(IV) chain-specific rat monoclonal antibodies against synthetic peptides reflecting sequences near the carboxy terminus of each noncollagenous (NC)1 domain. By Western blotting α6(IV) chain-specific antibody recognized 27-kD monomers and associated dimers of the human type IV collagen NC1 domain, which is the first demonstration of the presence in tissues of the α6(IV) polypeptide as predicted from its cDNA sequence. Immunofluorescence studies using anti-α6(IV) antibody demonstrated that in human adult kidney the α6(IV) chain was never detected in the glomerular basement membrane, whereas the basement membranes of the Bowman's capsules and distal tubules were positive. The staining pattern of the glomerular basement membrane was quite different from that obtained with the anti-α5(IV) peptide antibody. The α5(IV) and α6(IV) chains were colocalized in the basement membrane in the skin, smooth muscle cells, and adipocytes; however, little if any reaction was seen in basement membranes of cardiac muscles and hepatic sinusoidal endothelial cells. Thus, both genes are expressed in a tissue-specific manner, perhaps due to the unique function of the bidirectional promoter for both genes, which is presumably different from that for COL4A1 and COL4A2.