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: 6896811
• 负责人: Jonathan A Cooper
• 金额: $ 47.84万
• 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/6896811
• 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（Dab 2）基因通过未知机制抑制肿瘤生长。有2种Dab 2蛋白形式，p96和p67似乎是接头蛋白。一些衔接蛋白调节内吞作用，其他衔接蛋白在信号转导中起作用。一个令人兴奋的想法，我们将测试Dab 2是，一些蛋白质做这两个。我们的总体目标是确定Dab 2在体内功能的分子机制。我们发现p96而不是p67与内吞蛋白结合并定位于网格蛋白包被的小凹。此外，p96和p67都含有与脂蛋白受体的内吞信号相关的PTB/PID结构域，以及与非肌肉肌球蛋白（肌球蛋白VI）结合的单独结构域，该结构域与内吞作用有关。事实上，我们已经证明，肾脏缺乏Dab 2会导致蛋白质转运缺陷，类似于缺乏脂蛋白受体megalin的小鼠。因此，我们假设Dab 2正常调节megalin交通。此外，我们还发现Dab 2在原肠胚形成之前还有另一个重要的功能，此时p67是主要形式。Dab 2突变胚胎在胚外上皮（内脏内胚层）中存在缺陷，这种缺陷阻止了前后轴的诱导。该表型似乎是由TGF β家族成员Nodal的信号传导受损引起的。我们假设p67参与内脏内胚层的信号传导。由于p67缺乏包被小凹定位的信号，因此它可能直接参与信号转导。我们将确定Dab 2突变内脏内胚层中有缺陷的信号通路，并确定需要Dab 2的步骤。我们将测试p96是否专门用于调节肾脏转运，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。再加上在肾脏和内脏内胚层的Dab 2蛋白的亚细胞定位的研究，这种方法将提供证据是否Dab 2的胚胎需求是一个间接的后果，在受体运输的作用，或是否p67具有信号传导功能。我们将研究Dab 2调节肾脏中巨蛋白运输的机制，特别是与网格蛋白包被的小窝和肌球蛋白VI成分结合的重要性。我们还发现在丝裂原刺激的细胞中，Dab 2被MAP激酶磷酸化，并将探讨Dab 2磷酸化在调节生物反应中的意义。