The Sir2-p53-IGF link in mammalian life-span control

描述（由申请人提供）：我们已经产生了一种转基因小鼠，其中肿瘤抑制因子p53（p44）的短同种型的过度表达导致健康和生命的过早丧失。我们打算描绘的分子机制，增加p44的表达加速这个动物模型的正常衰老过程。在特定目的1中，我们检验了p44与p53比值的变化可以调节哺乳动物寿命的假设。在这个目标中，我们表征了小鼠和人类正常衰老过程中p44的表达（实验1.1），并使用细胞中的RNAi（实验1.2）和动物中的p44缺陷型p53等位基因（实验1.3）来操纵p44水平，以确定其在全长p53背景下与衰老速率的相关性。在SPECIFIC AIM 2中，我们检验了p53通过偶联Sir 2和IGF-1通路调节小鼠寿命的假设。为此，我们提出了一系列的生物化学实验来阐明p44可能干扰p53和Sir 2之间相互作用的分子机制，包括改变p53与其已知结合配偶体的相互作用（实验2.1）;改变p53翻译后修饰，包括磷酸化和乙酰化/脱乙酰化（实验2.2）;改变p53亚细胞定位（实验2.3）;以及干扰p53依赖性转录复合物的组装或功能（实验2.4）。如果p53是小鼠中Sir 2a和IGF信号传导之间的联系，那么p44将该联系解偶联，并且该联系如何正常发挥作用的线索在于为什么当p44过量时它不起作用。最后，在《特定目的3》中，我们检验了Igf-1受体是p44改变哺乳动物健康和寿命的主要下游效应子的假设。在这个目标中，我们利用小鼠遗传学的力量，以确定是否Igf-1受体，这是改变细胞和组织的p44纯合转基因小鼠，是介导的关键组成部分，在这个哺乳动物系统的健康和生命的损失。由于该基因是IGF-1信号转导中关键的第一步，因此本研究的重点是p53与信号通路的相互作用，该信号通路在低等生物（如C. elegans和D.黑腹菌

DESCRIPTION (provided by applicant): We have generated a transgenic mouse in which over-expression of the short isoform of the tumor suppressor p53 (p44) results in the premature loss of health and life. We intend to delineate the molecular mechanism by which increased p44 expression accelerates the process of normal aging in this animal model. In SPECIFIC AIM 1, we test the hypothesis that a change in the p44-to-p53 ratio can modulate mammalian longevity. In this Aim, we characterize the expression of p44 during normal aging in both mouse and human (Experiment 1.1) and determine its relevance in the context of full-length p53 to the rate of aging using RNAi in cells (Experiments 1.2) and a p44-deficient p53 allele in animals (Experiment 1.3) to manipulate p44 levels. In SPECIFIC AIM 2, we test the hypothesis that p53 modulates life-span in the mouse by coupling the Sir2 and IGF-1 pathways. In this Aim, we propose a series of biochemical experiments to elucidate the molecular mechanism by which p44 might interfere with the interaction between p53 and Sir2, including altering the interaction of p53 with its known binding partners (Experiment 2.1); altering p53 post- translational modifications, including phosphorylation and acetylation/deacetylation (Experiment 2.2); altering p53 sub-cellular localization (Experiment 2.3); and, interfering with the assembly or function of p53- dependent transcription complexes (Experiment 2.4). If p53 is the link between Sir2a and IGF signaling in the mouse, then p44 uncouples the link, and the clues to how this link functions normally lie in why it is non- functional when p44 is in excess. Finally, in SPECIFIC AIM 3, we test the hypothesis that the Igf-1 receptor is the principal downstream effector by which p44 alters mammalian health- and life-span. In this aim, we exploit the power of mouse genetics to determine if the Igf-1 receptor, which is altered in cells and tissues of p44 homozygous transgenic mice, is the key component mediating the loss of health and life in this mammalian system. Because this gene is the critical first step in IGF-1 signal transduction, this Aim focuses on the interaction of p53 and the signaling pathway that plays a major role in life-span determination in lower organisms, such as C. elegans and D. melanogaster.