An endothelial-specific switch in gene expression.

描述（由申请人提供）：近交系小鼠RIIIS/J是1型血管性血友病（VWD）的模型，VWD是人类最常见的出血性疾病。我们之前发现RIIIS/J低VWF的原因是编码n -乙酰半乳糖氨基转移酶GALGT2基因的调控突变Mvwfl。Mvwfl导致组织特异性转换，从大多数小鼠品系中观察到的肠上皮表达模式转换为血管内皮模式，导致VWF的异常翻译后修饰和加速清除。导致这一显著转变的确切顺式监管因素尚未被确定。我们假设，对RIIIS/J开关的调控元件的表征将显著促进我们对血管和肠道特异性基因表达程序的理解。该项目将利用直接序列分析和工程细菌人工染色体系统（BACs）来确定负责转换的基因组区域。不同Mvwfl小鼠品系的序列比较分析应确定共享Mvwfl单倍型片段的最小长度。嵌合RIIIS/J:C57BL6/J BAC将通过将RIIIS/J候选序列片段交换到已知具有肠上皮Galgt2表达的C57BL/6J BAC中来进行工程设计。将产生转基因小鼠并检测Galgt2表达的组织模式。赋予RIIIS/J Galgt2表达模式的构建体应该定义包含关键调控元件的区域，该区域将通过额外的BAC突变进一步表征。我们的发现将为内皮基因表达所需的顺式调控元件提供新的见解，对其他组织特异性基因表达程序和血管基因治疗具有潜在的广泛意义。

DESCRIPTION (provided by applicant): The inbred mouse strain RIIIS/J is a model for type 1 von Willebrand Disease (VWD), the most common bleeding disorder in humans. We previously found that the cause of low VWF in RIIIS/J is a regulatory mutation, Mvwfl, in the gene encoding an N-acetylgalactosaminyltransferase, GALGT2. Mvwfl causes a tissue-specific switch from the intestinal epithelial expression pattern observed in most mouse strains to a vascular endothelial pattern, resulting in aberrant post-translational modification of VWF and accelerated clearance. The precise cis-regulatory elements responsible for this remarkable switch have not yet been identified. We hypothesize that characterization of the regulatory element(s) responsible for the RIIIS/J switch will significantly advance our understanding of vascular and intestine specific gene expression programs. This project will utilize direct sequence analysis and engineered bacterial artificial chromosome systems (BACs) to identify the genomic regions responsible for the switch. Comparative sequence analysis of different Mvwfl mouse strains should determine the minimum length of the shared Mvwfl haplotype block. Chimeric RIIIS/J:C57BL6/J BACs will be engineered by swapping fragments of the RIIIS/J candidate sequence into a C57BL/6J BAC that is known to exhibit intestinal epithelial Galgt2 expression. Transgenic mice will be generated and tested for tissue patterns of Galgt2 expression. Constructs that confer the RIIIS/J Galgt2 expression pattern should define the region containing the critical regulatory element(s), which will be further characterized by additional BAC mutations. Our findings should provide new insight into the cis-regulatory elements required for endothelial gene expression, with potential broad implications for other tissue specific gene expression programs and vascular gene therapy.