Functional Analysis of Collagen IV by Gene Targeting

Col4a6 null mice were created by introducing Neo D1 R D1 gene into exon II of col4a6 gene。我们分析了如果基因被转录和翻译成α6（IV）多肽在位点杂交、北溴杂交和免疫性化学染色使用α链特定单克隆抗体。该结果证明了α6（IV）多肽并没有出现在任何非常规的组织或组织中。我们也分析了如果老鼠有某些形态，但我们没有检测到野生类型和col 4a 6 null老鼠之间的差异。我们预测了IV型聚合物的三种分子形式的存在; [α1（IV）]2α2（IV）， α3（IV）α4（IV）和[α5（IV）]2α6α（IV） by use of the monoclonal antibodies specific for α（IV） chains。[α1（IV）]2α2（IV）形式存在于所有基础薄膜中，其中α3（IV）α4（IV）α5（IV）形式被定位于凝胶和碱性基础薄膜和[α5（IV）]2α6（IV）形式被定位于Bowman's Capsules of肾脏和皮肤基础薄膜区域。三种分子形式的异构分布被标记为它们可能在不同的基础薄膜中具有特定的作用。To describe the biological roles of the molecules， we should think of how the three molecules are incorporated into the supramolecular aggregates of each basement membrane.We identified and characterized the breakpoint sequences of the deletion of DNA from a patient of diffuse leiomatosis associated with Alport syndrome。结果说明了删除删除的第一个编码前COL 4A 5和第二个COL 4A 6。他们还展示了断裂点分享相同的序列，使其更接近拓扑异构体I和II的共识。另一种DNA证据认为，男性患者是变异的物质。本研究对DL病理学及其伦理学的理解有很大的启示。

Col4a6 null mice were created by introducing NeoィイD1RィエD1 gene into exon II of col4a6 gene. We analyzed if the gene is transcribed and translated into α6(IV) polypeptide by in site hybridization, Northern-blot hybridization, and immunohistochemical staining using α chain-specific monoclonal antibodies. The results demonstrated that the α6(IV) polypeptide was not stained at all in any tissues or organs in col4a6 null mice. We also analyzed if the mice have some phenotypes, but we have not detected ant differences between wild type and col4a6 null mice so far.We predicted the presence of the three molecular forms of type IV collagen ; [α1(IV)]2α2(IV), α3(IV)α4(IV)α5(IV), and [α5(IV)]2α6α(IV) by use of double staining of the monoclonal antibodies specific for α(IV) chains. The [α1(IV)]2α2(IV) form is present in all basement membranes, whereas the α3(IV)α4(IV)α5(IV) form is localized in glomeruler and alveolar basement membranes and the [α5(IV)]2α6(IV) form is in the Bowman's capsules of the kidney and dermal basement membrane regions. Heterogeneous distribution of the three molecular forms indicated that they may have specific roles in different basement membranes. To describe the biological roles of the molecules, we should think of how the three molecules are incorporated into the supramolecular aggregates of each basement membrane.We identified and characterized the breakpoint sequences of the deletion of DNA from a patient of diffuse leiomyomatosis associated with Alport syndrome. The results demonstrated that a deletion eliminates the first coding exon of COL4A5 and the first two of COL4A6. They also showed that the breakpoints share the same sequence, which is in turn closely homologous to the consensuses of topoisomerases I and II. Additional DNA evidence suggested that the male patient is a somatic for the mutation. This study is greatly relevant to the understanding of DL pathogenesis and its etiology.