Characterization of Fortilin, A Novel Anti-p53 Protein

描述（由申请人提供）：动脉粥样硬化是主要原因 冠状动脉疾病、外周血管疾病和脑血管疾病 疾病在这个国家，折磨着数百万美国人。的发病机制 动脉粥样硬化是复杂的，但平滑肌增殖失调 cells细胞is involved牵连as an important重要proponent支持者.使用p53的遗传学研究 敲除小鼠表明，功能性053的存在是预防性的， 在高脂血症环境中对抗动脉粥样硬化。另外研究 已经表明p53抑制剂，如Mdm-2和IE84， 加速动脉粥样硬化我们的实验室发现了一种新的p53抑制剂， 将其命名为福尔替林。在我们的实验室中， 结果表明：（1）Fortilin的肽段序列在不同的氨基酸序列中高度保守， 不同的物种(2)Fortilin的信息在所有正常人中重新表达 组织，而其在癌细胞系中的表达更高。(3)福尔蒂安 表达在恶性组织中比在良性组织中更显著。(4)福尔蒂林 结合p53。(5)Fortilin的过度表达阻止了细胞 足叶乙甙诱导的细胞凋亡(6)Fortilin的过度表达 防止细胞经历由p53表达诱导的凋亡。（七） Fortilin的过表达抑制p53介导的BAX的反式激活， 促凋亡分子。(8)Fortilin的表达上调， 动脉粥样硬化。提出了三个主要的具体目标，以进一步表征 fortilin和fortilin-p53相互作用：目的1：确定 与p53和其他分子相互作用的fortilin。我们将首先确定 Fortilin的哪个结构域通过产生和 评价各种缺失突变体与福替林相互作用的能力 （目标1.1）。然后，我们将确定fortilin结构域的区域， 与p53相互作用所需的（Aim1.2）。最后，我们将建立 fortilin是否被sentrin和其他泛素样蛋白共价修饰 分子（目标1.3）。目的2：表征p53与p53相互作用的区域， 和其他分子。在这个主要目标中，我们确定哪个领域 (Aim 2.1）和p53的区域（Aim 2.2）参与福替林结合，使用 缺失突变体和点突变体。然后我们将评估 p53的fortilin结合区是p53与Mdm 2、Mdm-X相互作用所必需的 和BRCA 2、p53结合和抑制分子（Aim 2.3）。目标3： 探讨fortilin与p53相互作用对细胞凋亡的调控作用。我们 将确定福替林是否抑制p53介导的凋亡（Aim3.1）， p53介导的Bax反式激活（Aim 3.2）和p53结合至 基础反应元件（Aim 3.3）通过其直接结合p53。知识 我认为这一建议将使我们为未来打下坚实的基础 干预人类动脉粥样硬化。

DESCRIPTION (provided by the applicant): Atherosclerosis is the leading cause of coronary artery disease, peripheral vascular disease and cerebrovascular disease in this country, afflicting millions of Americans. The pathogenesis of atherosclerosis is complex, yet dysregulated proliferation of smooth muscle cells is implicated as an important proponent. A genetic study using p53 knockout mice suggested that he presence of functional 053 is preventive against atherosclerosis in a hyperlipidemic environment. In addition, studies have indicated that the presence of p53 inhibitors, such as Mdm-2 and IE84, accelerates atherosclerosis. Our laboratory identified a new p53 inhibitor and designated it as fortilin. Characterization of fortilin in our laboratory so far showed: (1) The peptide sequence of fortilin is highly conserved among different species. (2) The message of fortilin is resent in all normal human tissues while its expression is greater in cancerous cell lines. (3) Fortiin expression is more prominent in malignant than I benign tissue. (4) Fortilin binds p53. (5) The overexpression prevents of fortilin prevents cells from undergoing etoposide-induced apoptosis. (6) The over expression of fortilin prevents cells from undergoing apoptosis induced by the expression of p53. (7) The overexpression of fortilin inhibits p53-mediated transactivation of BAX, a pro-apoptotic molecule. (8) The expression of fortilin is up regulated in atheroma. There are three major Specific Aims proposed to further characterize fortilin and the fortilin-p53 interaction: Aim 1: To define the region of fortilin that interacts with p53 and other molecules. We will first determine which domain of fortilin participates in p53 binding by generating and evaluating various deletion mutants for their ability to interact with fortilin (Aim1.1). We will then identify the region of that fortilin domain that is required for the interaction with p53 (Aim1.2). Finally, we will establish whether fortilin is covalently modified by sentrin and other ubiquitin-like molecules (Aim 1.3). Aim 2: To characterize the region of p53 that interacts with fortilin and other molecules. In this major aim, we determine which domain (Aim 2.1) and region (Aim 2.2) of p53 participates in fortilin binding, using deletion and point mutants, respectively. We will then evaluate whether the fortilin-binding region of p53 is required for p53 to interact with Mdm2, Mdm-X and BRCA2, p53 binding and inhibitory molecules (Aim 2.3). Aim 3: To investigate the regulatory role of fortilin-p53 interaction in apoptosis. We will determine whether fortilin inhibits p53-mediatd apoptosis (Aim3.1), p53-mediated transactivation of Bax (Aim 3.2) and p53 binding to the Bas-responsive element (Aim 3.3) THROUGH ITS DIRECT binding of p53. Knowledge gained I this proposal will enable us to have a solid foundation for the future intervention of human atherosclerosis.