Type I osteogenesis imperfecta (01) is a dominantly inherited disease characterized clinically by bone fractures during childhood, blue sclerae, and frequent hearing loss accompanied by a decreased content of type I collagen in bone and skin.Cultured skdn fibroblasts from three individuals affected with the disease produce half-normal levels of type I procollagen, a disulfide-bonded trimer that contains two proal(I) chains and one proa2(I) chain. In normal cells, proal(I) and proa2(I) are synthesized in a 2:1 ratio and only assembled molecules are secreted. In contrast, the OI cells contain equimolar amounts of proal(I) and proa2(I), which suggests that trimer assembly and secretion are limited by the level ofproal(I) synthesis. The "extra" proa2(I) in the OI cells is in a nondisulfide-bonded configuration and is not secreted but apparently contributes to an increased level of intracellular degradation. Thus, decreased production oftype I procollagen in these patients is the result of decreased synthesis of proal(I). These results suggest that the stoichiometry of proa chains in type I procollagen is determined by the conformation of the chains rather than the ratio in which they are synthesized, that molecules containing more than a single proa2(I) chain are not assembled, and that the production of this heteropolymeric molecule may be effectively regulated by controlling the synthesis of only one of the subunits.Biochemical investigation of human hereditary disorders has provided valuable insight into mechanisms of gene expression. There are many examples ofmutations that alter the expression ofa single gene product, but mutations in regulatory genes that control more than one transcriptional unit are rare (1, 2). In type I osteogenesis imperfecta (01), a dominantly inherited disease that presents clinically with bone fragility, blue sclerae, and presenile hearing loss (3, 4), the production of type I collagen by cultured cells (5) and the content of type I collagen in skin (6, 7) and bone (8) is decreased by =50%. Because type I procollagen contains proal(I) and proa2(I) chains that are coded for by separate mRNA molecules (9), the primary defect in type I OI may involve a regulatory gene that controls the synthesis of proal(I) and proa2(I) chains coordinately.We have studied cultured dermal fibroblasts derived from three patients with type I OI and now report that decreased production of type I procollagen is caused by a decrease in the intracellular level of proal(I) chains. Consequently, there is a relative excess ofproa2(I) chains that do not assemble into multimers but are degraded intracellularly instead. These findings suggest a model for controlling the production. of type I procollagen (or other heteropolymers) by regulating the production of a single subunit.
MATERIALS AND METHODSCell Culture. Skin fibroblasts were obtained by dermal biopsy and explant growth. Cells were maintained as described (10) described (10) with the following exceptions. In experiments designed to measure intracel...
A B S T R A C T Dermal fibroblasts in culture from a woman with a mild to moderate form of osteogenesis imperfecta synthesize two species of the proa2-chain of type I procollagen. One chain is normal. The abnormal chain has a slightly faster mobility than normal during electrophoresis in sodium dodecyl sulfate polyacrylamide gels. Analysis of cyanogen bromide peptides of the proa-chain, the a-chain, and of the mammalian collagenase cleavage products of the proa-and a-chains indicates that the abnormality is confined to the a2(I)CB4 fragment and is consistent with loss of a short triple-helical segment. Type I collagen production was decreased, perhaps because the molecules that contained the abnormal chain were unstable, with a resultant alteration in the ratio of type III to type I collagen secreted into culture medium. Collagen fibrils in bone and skin had a normal periodicity but their diameters were 50% of control; the bone matrix was undermineralized. The structural abnormality in the a2(I)-chain in this patient may affect molecular stability, intermolecular interactions, and collagenmineral relationships that act to decrease the collagen content of tissues and affect the mineralization of bone.
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