MOLECULAR BASIS OF OSTEOGENESIS IMPERFECTA

成骨不全(OI)是一种临床、遗传和生化方面的疾病 以骨脆性和其他结缔组织为特征的异质性疾病 组织异常。在绝大多数受影响的人中， 临床症状由两个基因(COL1A1和COL1A1)突变引起 COL1A2)，编码I型前胶原的链。的目标 本申请中提出的研究是为了描述突变的特征 在导致不同形式的OI的胶原基因中，确定 突变的性质与其所在基因之间的关系 定位和临床表型，以确定突变如何改变 含有异常分子的胞内和胞外行为 链，以确定新的显性突变的起源的亲本，并 确定胶原基因突变的嵌合体程度 最先受影响的人的父母。对于改变链的突变 结构，突变的位置将由多肽近似。 映射，突变的准确位置将由 单碱基错配化学裂解或单链构象 多态，成纤维细胞或基因组中cDNAs的合适区域 DNA将被扩增并直接测序或克隆到M13和 将对选定的克隆进行排序。对于改变表达的突变 在胶原基因中，异常等位基因将通过缺乏 从该基因合成的mRNA中表达的多态标记(在 杂合子)和突变的结构被鉴定和 用碱基错配化学方法搜索基因后的特征 单链构象多态的裂解检测区域 差异之处。I型前胶原蛋白中链异常的作用 分子将通过检查分泌物的动力学和 粗面内质网结合蛋白对细胞分泌、分泌效率的影响 具有一条或多条异常链的分子的分泌， 在EM水平上，这些分子对分子结构的影响 分子的热稳定性，以及分子参与的能力 在纤维形成过程中。最后，马赛克的程度将取决于 对父母的组织进行检查并测量是否存在 用适当的限制性内切酶消化或通过 等位基因特异性寡核苷酸杂交。这些研究应该会有所帮助 为了了解OI的分子基础，以及突变是如何翻译的 到表型。最后，它们对多发性硬化症的发病机制有一定的影响。 常见的骨形成障碍。

Osteogenesis imperfecta (OI) is a clinically, genetically and biochemically heterogeneous disorder characterized by bone fragility and other connective tissue abnormalities. In the vast majority of affected individuals the clinical condition results from mutations in the two genes (COL1A1 and COL1A2) that encode the chains of type I procoliagen. The objectives of the studies proposed in this application are to characterize the mutations in the collagen genes that give rise to different forms of OI, to determine the relationship among the nature of the mutation, the gene in which it is located, and the clinical phenotype, to determine how mutations alter the intracellular and extracellular behavior of molecules that contain abnormal chains, to identify the parent of origin of new dominant mutations, and to determine the extent of mosaicism for mutations in collagen genes among parents of first affected individuals. For mutations that alter chain structure, the site of the mutation will be approximated by peptide mapping, the precise location of the mutation will be determined by single-base mismatch chemical cleavage or single stranded conformational polymorphisms, the appropriate region of cDNA from fibroblasts or genomic DNA will be amplified and either sequenced directly or cloned into M13 and selected clones will be sequenced. For mutations that alter the expression of collagen genes, the abnormal allele will be identified by the absence of expressed polymorphic markers from the mRNA synthesized from that gene (in heterozygotes) and the structure of the mutation identified and characterized following search of the gene using base mismatch chemical cleavage of single stranded conformational polymorphisms to detect regions of difference. The effects of abnormal chains in type I procollagen molecules will be determined by examinining the kinetics of secretion and the effects of RER binding proteins on secretion, the efficiency of secretion of molecules with one or more than one abnormal chain, the effect-,of these molecules on molecular structure at the EM level, the thermal stability of molecules, and the ability of molecules to participate in fibrillogenesis. Finally, the extent of mosaicism will be determined by examination of tissues from parents and measuring the presence of the mutant allele by digestions with appropriate restriction enzymes or by allele-specific oligonucleotide hybridization. These studies should help to understand the molecular basis of OI, and how mutations are translated to phenotype. Finally, they have implications for the pathogenesis of more common disorders of bone formation.