MOLECULAR MECHANISMS OF RENAL INJURY

基底膜由IV型胶原蛋白、层粘连蛋白， 蛋白聚糖和巢蛋白。IV型胶原蛋白由六个家族组成 不同的α链肾小球基底膜（GBM）胶原， 例如，在两相中形成α-3（IV）和α-5（IV）， 胶原蛋白出现较晚，最终在最初的原聚体中占主导地位 由α-1（IV）和α-2（IV）链形成。基底膜 相反，来自肝脏的，通常不表达α-5（IV）或 α-6（IV）胶原蛋白在其初始形成后。 这种从胎儿α-1（IV）和α-2（IV）胶原蛋白向α- 人GBM中的α-3（IV）、α-4（IV）和α-5（IV）链是一种 促进毛细血管环形成的发育事件， 肾小球我们相信这种发育转变改善了 毛细过滤器的结构完整性，通过保护性地增强其 对肾小球滤过部位的局部蛋白酶的抗性。这 X连锁Alport患者的大多数肾脏中存在转换缺陷 综合征（XAS）谁有突变的α-5（IV）基因，并继续 到肾衰竭 我们建议通过以下方法来研究这种发育转换的机制： 研究alpha-5（IV）胶原基因的超分子控制。 我们假设，开关是门控的染色质开放， 选择的组织，并在DNA结合的模块化控制下， proteins.确定这些基因座控制区（LCR）的坐标 是理解这一监管的更大方面的第一步 过程 对IV型胶原蛋白转换重要的调节区域将 通过绘制肾脏中独特的DNA酶1超敏位点来研究 远离人类α-5（IV）基因起始的染色质。我们有 从该区域获得含有基因组DNA的BAC克隆， 染色体Xq22.2，并制定了一项战略， 转基因小鼠中的人α-5（IV）转录物。候选 超敏位点将表达为转基因， 功能活动、转换、位置独立性和拷贝数- 依赖然后，识别为LCR的超敏位点可以 在随后的二级核小体水平上进行了探索 问题研究

Basement membranes are composed of type IV collagen, laminins, proteoglycans and entactin. Type IV collagen comprises a family of six different alpha-chains. Glomerular basement membrane (GBM) collagen, for example, is formed in two phases with alpha-3(IV), and alpha-5(IV) collagens appearing later and eventually dominating protomers initially formed from alpha-1 (IV) and alpha-2(IV) chains. Basement membranes from liver, in contradistinction, do not normally express alpha-5(IV) or alpha-6(IV) collagens after their initial formation. This switch from fetal alpha-1 (IV) and alpha-2(IV) collagens to alpha- 3(IV), alpha-4(IV), and alpha-5(IV) chains in human GBM is a developmental event that advances the fabrication of capillary loops in the glomerulus. We believe this developmental switch improves the structural integrity of the capillary filter by protectively enhancing its resistance to local proteases at the site of glomerular filtration. This switch is defective in most kidneys from patients with x-linked Alport Syndrome (XAS) who have mutations in the alpha-5(IV) gene and go on to renal failure. We propose to examine the mechanism of this developmental switch by studying the supramolecular control of the alpha-5(IV) collagen gene. We hypothesize that switching is gated by the opening of chromatin in selected tissues, and is under the modular control of DNA-binding proteins. Determining coordinates for these locus control regions (LCRs) is the first step in understanding the larger aspects of this regulatory process. Regulatory regions important for the switching of type IV collagen will be studied by mapping unique DNasel hypersensitive sites in kidney chromatin remote to the start of the human alpha-5(IV) gene. We have obtained a BAC clone containing genomic DNA from this region on chromosome Xq22.2, and have developed a strategy for measuring human alpha-5(IV) transcripts in transgenic mice. Candidate hypersensitive sites will be expressed as transgenes and evaluated for functional activity, switching, position-independence and copy number- dependence. Hypersensitive sites identified as LCRs can then e explored at the level of the nucleosome in subsequent, second-order studies.