Functional Studies of VHL by Conditional Gene Targeting

通过条件基因靶向进行 VHL 功能研究

• 批准号: 6633430
• 负责人: Volker Hans Haase
• 金额: $ 7.96万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-06-15 至 2004-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6633430
• 关键词: Von Hippel Lindau syndrome; apoptosis; cell growth regulation; disease /disorder model; gene mutation; gene targeting; genetically modified animals; hypoxia inducible factor 1; laboratory mouse; microarray technology; molecular pathology; renal tubule

DESCRIPTION (provided by applicant): Patients with mutations in the von Hippel-Lindau gene (VHL) develop a pleomorphic familial tumor syndrome that is characterized by the development of highly vascularized tumors with different biological behaviors such as CNS hemangioblastomas and clear cell renal cell cancers (RCC) which are often preceded by renal cysts. Recently the VHL protein (pVHL) has been found to regulate the stability of hypoxia-inducible-factor (HIF). However not all aspects of this disease, such as the malignant phenotype of RCCs, can be easily explained by this interaction. A VHL mouse model would provide a powerful tool for the identification of molecular factors that are critical for the pathogenesis of the VHL phenotype. However, the development of such a model was hampered by the fact that conventional VHL knockout mice died during midgestation from placental failure, which initially precluded a functional analysis of this gene in the adult. In order to overcome this problem with embryonic lethality, the Principal Investigator has generated mice that allow tissue-specific deletions of Vhl in the adult through Cre/loxP-mediated recombination. Several conditional mouse mutants have been generated. Inactivation of Vhl in the liver for example leads to the accumulation of neutral lipids in hepatocytes, proliferation of endothelial cells, as well as erythrocytosis, all of which is consistent with pVHL's role in the regulation of HIF. At the moment it remains unclear whether the phenotypes that were observed in conditional VHL knockout mice can be solely attributed to the up-regulation of HIF-1 dependent target genes and whether pVHL controlled molecular pathways exist that are not regulated by HIF. The studies proposed here will address these questions and aim at defining the critical molecular mediators of the VHL phenotype by generating several tissue-specific conditional mouse mutants that are double deficient in Vhl and Hif-1. The molecular analysis of these mutants with microarrays might provide important insights into the regulation of growth and apoptosis of renal tubule cells. The proposed investigations represent a continuation and expansion of research that was initially described in the applicant's K08 and is based on data that were generated under this grant. They will be carried out in the laboratory of Dr. Volker H. Haase, who is an Assistant Professor of Medicine in the University of Pennsylvania's Renal Division. Dr. Michael Madaio, Professor of Medicine, will function as Dr. Haase's mentor for this grant.

描述(由申请人提供)： 具有von Hippel-Lindau基因(VHL)突变的患者会出现多形性家族性肿瘤综合征，其特征是发生具有不同生物学行为的高度血管化的肿瘤，如中枢神经系统血管母细胞瘤和肾透明细胞癌(RCC)，这些肿瘤通常先于肾囊肿。最近发现VHL蛋白(PVHL)调节缺氧诱导因子(HIF)的稳定性。然而，这种相互作用并不能很容易地解释这种疾病的所有方面，例如RCC的恶性表型。VHL小鼠模型将为鉴定与VHL表型发病密切相关的分子因子提供有力的工具。然而，由于传统的VHL基因敲除小鼠在妊娠中期死于胎盘衰竭，这一事实阻碍了这种模型的发展，这最初排除了对该基因在成人中的功能分析。为了克服胚胎致死性的这一问题，首席研究员培育了一只小鼠，通过Cre/loxP介导的重组，在成年鼠中允许组织特异性删除VHL。已经产生了几个有条件的小鼠突变体。例如，VHL在肝脏中的失活导致中性脂在肝细胞中积聚，内皮细胞增殖，以及红细胞增多，所有这些都与pVHL在调节HIF中的作用是一致的。目前尚不清楚在条件性VHL基因敲除小鼠中观察到的表型是否完全归因于HIF-1依赖的靶基因的上调，以及pVHL是否存在不受HIF调控的分子通路。本文提出的研究将解决这些问题，并旨在通过产生几个VHL和Hif-1双重缺陷的组织特异性条件突变来确定VHL表型的关键分子介质。利用基因芯片对这些突变体进行分子分析，可能为调控肾小管细胞的生长和凋亡提供重要的信息。拟议的调查代表了最初在申请人的K08中描述的研究的继续和扩展，并基于根据这一赠款产生的数据。他们将在宾夕法尼亚大学肾科医学助理教授沃尔克·H·哈斯博士的实验室进行。医学教授迈克尔·马代奥博士将担任哈斯博士的这笔赠款的导师。