MOLECULAR BIOLOGY OF COLLAGEN TYPES V AND VII

描述：本提案包括V型胶原蛋白及其 在人类疾病中的作用，以及对组织特异性表达 VII型胶原基因 研究将包括：1）类型V的作用 Ehlers-Danlos综合征（EDS）中的胶原蛋白。 分子缺陷还有待于 在人类的V型胶原蛋白中有报道。 然而，一个患有EDS的病人， 伊藤黑色素减少症有一个平衡的（X;9）易位，似乎有 α 1（V）胶原基因（COL 5A 1）内的染色体断裂点。 的 将确定COL 5A 1内重排的性质，以提供 深入了解易位机制和分子基础的 这位EDS患者的表型。 在一项更深远的研究中， 将开发用于基于PCR的V型突变扫描的探针 EDS患者的胶原DNA和RNA序列。 在这些领域开发的探针 研究将用于调查一种独特的临床资源，包括RNA和 来自590名EDS患者的基因组DNA，其中I型和III型胶原蛋白是 正常 在这些研究中开发的探针和策略将有助于 其他研究遗传性结缔组织疾病的研究人员。 （二） 前-α 3（V）胶原蛋白链的表征。 Pro-a3（V）是唯一的 V型胶原蛋白链，其完整的一级结构和核 酸性探针不可用。 将分离全长cDNA序列， 提供pro-a3（V）结构，并提供用于详细分析的探针， 图10示出了pro-a3（V）RNA的组织分布的表征。 严格 通过以下方法检查各种V/Xi型胶原蛋白链的共表达： 胚胎和成年小鼠组织的原位杂交应该提供 深入了解V/Xi链的各种组合在 发育和体内平衡。 a3（V）基因将在小鼠中定位， 人类，以确定它是否与遗传基因座共定位 异常 将开发探针，以测定以下中的pro-a3（V）缺陷： 上述EDS患者组。 3)转录控制 VII型胶原基因COL 7A 1。 高度自治的机制 将探索COL 7A 1的组织特异性表达。 研究将包括 在细胞培养物和转基因小鼠中的功能测定， 通过突变分析表征顺式作用元件， 结合特定的核因子。 应提供新的见解， 分化和组织特异性表达是在某些 伊特莉亚 COL 7A 1顺式作用元件也有可能帮助实现 基因治疗中医学上有用的基因的真皮特异性表达。

DESCRIPTION: This proposal comprises studies of type V collagen and its roles in human diseases and studies into the tissue-specific expression of the type VII collagen gene. Studies will include: 1) The role of type V collagen in Ehlers-Danlos syndrome (EDS). Molecular defects have yet to be reported in the type V collagen of humans. However, a patient with EDS and hypomelanosis of Ito has a balanced (X;9) translocation and appears to have a chromosomal breakpoint within the alpha 1(V) collagen gene (COL5A1). The nature of the rearrangement within COL5A1 will be determined to provide insights into translocation mechanisms and into the molecular basis of the phenotype in this EDS patient. In a more far-reaching study, molecular probes will be developed for PCR-based scanning for mutations in type V collagen DNA and RNA sequences of EDS patients. Probes developed in these studies will be used to survey a unique clinical resource comprising RNA and genomic DNA from 590 EDS patients in whom collagen types I and III are normal. Probes and strategies developed in these studies will be of use to other researchers studying heritable connective tissue disorders. 2) Characterization of the pro-a3(V) collagen chain. Pro-a3(V) is the only type V collagen chain for which a complete primary structure and nucleic acid probes are not available. Full-length cDNA sequences will be isolated, providing the pro-a3(V) structure and providing probes for detailed characterization of the tissue distribution of pro-a3(V) RNA. Rigorous examination of the co-expression of the various collagen type V/XI chains by in situ hybridization of fetal and adult mouse tissues should provide insights into roles of the various combinations of V/XI chains in development and homeostasis. The a3(V) gene will be mapped in mice and humans to determine whether it colocalizes with loci for heritable abnormalities. Probes will be developed to assay for pro-a3(V) defects in the panel of EDS patients described above. 3) Transcription controls of type VII collagen gene, COL7A1. Mechanisms governing the highly tissue-specific expression of COL7A1 will be explored. Studies will include functional assays in cell culture and transgenic mice and further characterization of cis-acting elements by mutation analyses and for ability to bind specific nuclear factors. New insights should be provided into how differentiation- and tissue-specific expression is achieved in certain ethelia. COL7A1 cis-acting elements also have potential to help achieve edermis-specific expression of medically useful genes in gene therapy.