Identification and Characterization of the Mouse PPCD1 Gene

小鼠 PPCD1 基因的鉴定和表征

基本信息

批准号：
8309049
负责人：
CHRISTOPHER A BRADFIELD
金额：
$ 33.86万
依托单位：
UNIVERSITY OF WISCONSIN-MADISON
依托单位国家：
美国
项目类别：
财政年份：
2011
资助国家：
美国
起止时间：
2011-08-01 至 2015-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8309049
关键词：
Address Alleles Animals Anterior Biochemical Biological Assay Blindness Candidate Disease Gene Cells Characteristics Chromosomes, Human, Pair 2 Chromosomes, Human, Pair 20 Collaborations Complex Cornea Corneal Endothelium Corneal Neovascularization Corneal Opacity Corneal dystrophy Cytokeratin DNA DNA Sequence Rearrangement Data Development Disease Endothelial Cells Exhibits Eye Family member Gene Expression Gene Expression Profile Generations Genes Genetic Glaucoma Goals Heterogeneity Heterozygote Human Human Chromosomes In Vitro Inheritance Patterns Inherited Iris Diseases Laboratories Light Maps Metaplasia Methods Michigan Molecular Mus Mutation Orthologous Gene Pathogenesis Patients Pattern Penetrance Phenotype Photoreceptors Positioning Attribute Proteins Pseudogenes Recombinants Regulation Retinal Retinal Ganglion Cells Role Sampling Sampling Studies Screening procedure Secondary to Signal Pathway Structure Syndrome System Testing Transcript Universities XCL1 gene anterior chamber base cell type chromatin immunoprecipitation design genetic linkage analysis histone acetyltransferase human disease in vitro Assay insight mouse model promoter

项目摘要

PROJECT SUMMARY/ABSTRACT The PPCD1 mouse arose from a spontaneous mutation in our mouse colony and exhibits an enlarged anterior chamber secondary to metaplasia of the corneal endothelium and blockage of the iridocorneal angle by the epithelialized corneal endothelial cells. The presence of stratified multilayered corneal endothelial cells with abnormal patterns of cytokeratin expression are remarkably similar to those observed in human corneal endothelial dystrophies, notably posterior polymorphous dystrophy (PPD), and the sporadic condition, iridocorneal endothelial syndrome (ICE). Secondary phenotypes observed in PPCD1 mice include corneal neovascularization, retinal ganglion cell loss, and photoreceptor loss, all significant causes of blindness in humans. The mouse PPCD1 phenotype exhibits an autosomal dominant pattern of inheritance, with complete penetrance on the sensitive DBA/2J background and significantly decreased penetrance on the C57BL/6J background. The objective of this proposal is to provide a molecular explanation for the murine PPCD1 phenotypes with hopes of shedding light on the human disease. We have mapped the mouse PPCD1 gene, designated "Ppcd1", to a 6.2 Mbp interval on Chromosome 2, and identified a hemizygous 87,000 bp duplication in this interval. The endpoints of the duplication are located in positions which disrupt the genes Csrp2bp and 6330439K17Rik. LOC100043552, a pseudogene located in the center of the sequence, is also presumably duplicated. Our primary candidate for the Ppcd1 gene is Csrp2bp. This prediction is based on decreased Csrp2bp expression levels in PPCD1 animals, preliminary evidence for abnormal eye size and corneal opacities in induced null alleles of Csrp2bp and the known physical interaction of this gene product with the product of the Zeb1 locius (the other known hereditary cause of PPCD in humans). We propose to confirm our hypothesis that mutation of Csrp2bp is responsible for the PPCD1 phenotype by replicating corneal endothelial cell metaplasia in independent recombinant mouse models. We propose to identify mechanisms by which Csrp2bp interactions with Zeb1 cause the PPCD1 phenotype. We will determine the temporal and spatial localization of normal and abnormal Csrp2bp gene expression, identify Csrp2bp-regulated gene promoters and determine if they are also regulated by Zeb1. We will develop methods to test the function and regulation of Csrp2bp and demonstrate how disruption of its function produces the PPCD1 phenotype. We will extend our findings to human PPCD through screening for mutations in CSRP2BP and the orthologs of 6330439K17Rik and LOC100043552, respectively, C20ORF12, and ZNF 133 in PPCD1 patients in whom ZEB1 has been excluded as a causative gene. Putative causative mutations will be tested using in vitro functional assays. Finally, through linkage analysis, we will look for C57BL/6J modifier loci that influence the penetrance of PPCD1.

项目摘要/摘要 PPCD1小鼠来自我们小鼠菌落的自发突变，并表现出肿大继发于角膜内皮转移的前室和虹膜内部的阻塞上皮角膜内皮细胞的角度。分层多层角膜的存在细胞角蛋白表达异常模式的内皮细胞与观察到的细胞非常相似在人角膜内皮营养不良中，尤其是后多态营养不良（PPD）和零星状态，虹膜核内皮综合征（ICE）。在PPCD1中观察到的次级表型小鼠包括角膜新血管形成，视网膜神经节细胞损失和感光器损失，所有这些都显着人类失明的原因。小鼠PPCD1表型表现出一种常染色体显性模式继承，在敏感的DBA/2J背景上完全渗透，并显着降低 C57BL/6J背景上的渗透率。该建议的目的是提供分子对鼠PPCD1表型的解释，希望阐明人类疾病。我们已将指定为“ PPCD1”的小鼠PPCD1基因映射到染色体上的6.2 MBP间隔，并且在此时间间隔内确定了半合87,000 bp的重复。重复的终点位于在破坏基因CSRP2BP和6330439K17RIK的位置。 LOC100043552，位于在序列的中心，也可能是重复的。我们的PPCD1基因的主要候选者是 CSRP2BP。该预测基于PPCD1动物的CSRP2BP表达水平降低，初步 CSRP2BP诱导的无效等位基因中的眼睛大小和角膜泥泞异常的证据该基因产物与Zeb1 locius的产物的物理相互作用（另一个已知的遗传人类PPCD的原因）。我们建议确认我们的假设是CSRP2BP的突变是负责的通过在独立重组中复制角膜内皮细胞化生的PPCD1表型鼠标模型。我们建议确定CSRP2BP与Zeb1相互作用引起的机制 PPCD1表型。我们将确定正常和异常的时间和空间定位 CSRP2BP基因表达，识别CSRP2BP调节的基因启动子，并确定它们是否也是由Zeb1调节。我们将开发测试CSRP2BP功能和调节的方法并证明其功能的破坏如何产生PPCD1表型。我们将把发现扩展到人类PPCD 通过筛选CSRP2BP和6330439K17RIK和LOC100043552的突变，在PPCD1患者中，C20ORF12和ZNF 133分别被排除为Zeb1的患者致病基因。推定的致病突变将使用体外功能测定进行测试。最后，通过链接分析，我们将寻找影响PPCD1渗透率的C57BL/6J修饰基因座。