In vivo functions of possible tumor suppressor Dab2

可能的肿瘤抑制因子 Dab2 的体内功能

• 批准号: 6752904
• 负责人: Jonathan A Cooper
• 金额: $ 47.56万
• 依托单位: FRED HUTCHINSON CANCER RESEARCH CENTER
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-06-01 至 2007-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6752904
• 关键词: SDS polyacrylamide gel electrophoresis; biological signal transduction; embryogenesis; endocytosis; endoderm; gene targeting; genetically modified animals; immunocytochemistry; kidney cell; laboratory mouse; mitogen activated protein kinase; myosins; phosphorylation; polymerase chain reaction; protein structure function; protein transport; renal tubule; southern blotting; tissue /cell culture; tumor suppressor genes; western blottings

DESCRIPTION (provided by applicant): The Disabled-2 (Dab2) gene inhibits tumor growth through unknown mechanisms. There are 2 Dab2 protein forms, p96 and p67 that appear to be adaptor proteins. Some adaptor proteins regulate endocytosis and others function in signal transduction. One exciting idea that we'll test for Dab2 is that some proteins do both. Our overall goal is to define the molecular mechanisms underlying the in vivo functions of Dab2. We've found that p96 but not p67 binds to endocytic proteins and localizes to clathrin-coated pits. In addition, both p96 and p67 contain a PTB/PID domain that associates with the endocytosis signals of lipoprotein receptors, and a separate domain that binds to a non-muscle myosin, myosin VI, that has been implicated in endocytosis. Indeed, we've shown that lack of Dab2 in the kidney causes protein transport defects similar to those of mice lacking the lipoprotein receptor, megalin. Thus we hypothesize that Dab2 normally regulates megalin traffic. In addition, we've found that Dab2 has another important function prior to gastrulating, when p67 is the predominant form. Dab2 mutant embryos have a defect in an extraembryonic epithelium, the visceral endoderm, and this defect prevents induction of the anterior posterior axis. The phenotype appears to result from impaired signaling by Nodal, a TGFbeta family member. We hypothesize that p67 is involved in signaling in the visceral endoderm. Since p67 lacks signals for coated pit localization, it is possible it is directly involved in signal transduction. We will identify the defective signaling pathways in Dab2-mutant visceral endoderm, and identify the step requiring Dab2. We will test whether p96 is specialized to regulate kidney transport and p67 is specialized for signal transduction, or whether each form is multifunctional. Coupled with studies on subcellular localization of Dab2 proteins in kidney and visceral endoderm, this approach will provide evidence whether the embryonic requirement for Dab2 is an indirect consequence of a role in receptor trafficking, or whether p67 has a signaling function. We will investigate the mechanism by which Dab2 regulates megalin traffic in the kidney, especially the importance of binding to components of clathrin-coated pits and myosin VI. We've also found that Dab2 is phosphorylated in mitogen-stimulated cells by MAP kinase, and will investigate the significance of Dab2 phosphorylation in regulating biological responses.

描述（由申请人提供）：Disabled-2 （Dab2）基因通过未知机制抑制肿瘤生长。有2种Dab2蛋白形式，p96和p67似乎是适配蛋白。一些接头蛋白调节胞吞作用，另一些在信号转导中起作用。我们将测试Dab2的一个令人兴奋的想法是，一些蛋白质同时具有这两种功能。我们的总体目标是确定Dab2在体内功能的分子机制。我们发现p96而不是p67结合内吞蛋白并定位于网格蛋白包被的凹坑。此外，p96和p67都含有一个与脂蛋白受体内吞信号相关的PTB/PID结构域，以及一个与内吞作用有关的非肌肉肌球蛋白（myosin VI）结合的单独结构域。事实上，我们已经证明，肾脏中缺乏Dab2会导致蛋白质运输缺陷，类似于缺乏脂蛋白受体meggalin的小鼠。因此，我们假设Dab2通常调节meggalin流量。此外，我们发现Dab2在原肠泌乳之前还有另一个重要的功能，当p67是主要形式时。Dab2突变胚胎在胚胎外上皮（内脏内胚层）有缺陷，这种缺陷阻止了前后轴的诱导。这种表型似乎是由TGFbeta家族成员Nodal的信号传导受损引起的。我们假设p67参与了内脏内胚层的信号传导。由于p67缺乏涂层坑定位的信号，它可能直接参与信号转导。我们将确定Dab2突变的内脏内胚层中有缺陷的信号通路，并确定需要Dab2的步骤。我们将测试p96是否专门用于调节肾脏运输，p67是否专门用于信号转导，或者每种形式是否都是多功能的。结合对Dab2蛋白在肾脏和内脏内胚层亚细胞定位的研究，该方法将提供证据，证明胚胎对Dab2的需求是参与受体运输的间接结果，还是p67具有信号功能。我们将研究Dab2在肾脏中调节巨噬细胞蛋白运输的机制，特别是与网格蛋白包被凹坑和肌球蛋白VI结合的重要性。我们还发现Dab2在有丝分裂原刺激的细胞中被MAP激酶磷酸化，并将研究Dab2磷酸化在调节生物反应中的意义。