TYPE-V COLLAGEN - MOLECULAR-STRUCTURE AND FIBRILLAR ORGANIZATION OF THE CHICKEN ALPHA-1(V) NH2-TERMINAL DOMAIN, A PUTATIVE REGULATOR OF CORNEAL FIBRILLOGENESIS

Previous work from our laboratories has demonstrated that: (a) the str iated collagen fibrils of the corneal stroma are heterotypic structure s composed of type V collagen molecules coassembled along with those o f type I collagen, (b) the high content of type V collagen within the corneal collagen fibrils is one factor responsible for the small, unif orm fibrillar diameter (25 nm) characteristic of this tissue, (c) the completely processed form of type V collagen found within tissues reta ins a large noncollagenous region, termed the NH2-terminal domain, at the amino end of its al chain, and (d) the NH2-terminal domain may con tain at least some of the information for the observed regulation of f ibril diameters. In the present investigation we have employed polyclo nal antibodies against the retained NH2-terminal domain of the alpha1( V) chain for immunohistochemical studies of embryonic avian corneas an d for immunoscreening a chicken cDNA library. When combined with cDNA sequencing and molecular rotary shadowing, these approaches provide in formation on the molecular structure of the retained NH2-terminal doma in as well as how this domain might function in the regulation of fibr illar structure. In immunofluorescence and immunoelectron microscopy a nalyses, the antibodies against the NH2-terminal domain react with typ e V molecules present within mature heterotypic fibrils of the corneal stroma. Thus, epitopes within at least a portion of this domain are e xposed on the fibril surface. This is in marked contrast to mAbs which we have previously characterized as being directed against epitopes l ocated in the major triple helical domain of the type V molecule. The helical epitopes recognized by these antibodies are antigenically mask ed on type V molecules that have been assembled into fibrils. Sequenci ng of the isolated cDNA clones has provided the conceptual amino acid sequence of the entire amino end of the alpha1(V) procollagen chain. T he sequence shows the location of what appear to be potential propepti dase cleavage sites. One of these, if preferentially used during proce ssing of the type V procollagen molecule, can provide an explanation f or the retention of the NH2-terminal domain in the completely processe d molecule. The sequencing data also suggest that the NH2-terminal dom ain consists of several regions, providing a structure which fits well with that of the completely processed type V molecule as visualized b y rotary shadowing.