Insoluble protein carbonyl: A critical determinant for mammalian longevity

描述(由申请人提供)：现在有大量文献证明，从无脊椎动物(如酵母和果蝇)到人类，蛋白质翻译后的变化，特别是蛋白质的甲基化，随着年龄的增长而发生。如果不被蛋白酶体降解，随着时间的推移，碳化的蛋白质会积累形成高分子量的不溶聚集体。不可溶聚集体的积累已被证明对细胞具有极大的毒性，并与几种与年龄有关的疾病，特别是神经退行性疾病有关。最近在我的实验室对各种短期(实验室小鼠、野生捕获的小鼠、大鼠)和长寿(裸鼠、蝙蝠和绒猴)哺乳动物物种的研究，包括近亲繁殖的自由繁殖和饮食(DR)限制的C57BL/6幼鼠和老年小鼠，首次显示出一个意想不到的普遍观察结果，即长寿物种比短寿物种具有更低水平的不溶性细胞蛋白羰基。这一数据有力地表明，寿命长的物种似乎比寿命短的物种具有更好的能力来减少不溶性蛋白质羰基的积累。由于一般情况下，蛋白质被蛋白质酶体标记为蛋白质降解，我们推测活跃的蛋白酶体功能可能是长寿物种用来维持良好蛋白质质量的防御机制之一。我们对啮齿动物物种的初步研究发现，活跃的蛋白酶体功能与延长寿命之间存在直接联系。综上所述，这项拟议的研究将扩大这些有趣的初步观察结果，以评估一般假设，即长寿物种表现出不溶蛋白质羰基的积累减少，并通过激活蛋白酶体机械来维持细胞活力。在这项提议中，我们希望使用各种非传统的短寿命和长寿命的成纤维细胞来检验这一假设。这个项目是第一个严格评估蛋白质羰化和蛋白酶体功能之间的串扰的研究，蛋白酶体功能在保护细胞免受广泛哺乳动物的蛋白质毒性方面发挥着关键作用。这些数据将使我们能够确定降低蛋白质毒性是否是进化过程中用来延长寿命的一种常见/公共机制。这项提议将通过追求以下具体目标来检验这一假设。具体目的1.验证有效的蛋白酶体功能反应、减少不溶蛋白的羰基和恢复细胞活力是长寿种成纤维细胞对氧化应激反应的共同特征的假设。具体目的2.验证蛋白酶体功能失活引发难溶蛋白羰基积累，从而导致长寿物种成纤维细胞死亡增加的假说。

DESCRIPTION (provided by applicant): It is now well documented in literature that post-translational changes in protein, particularly protein carbonylation, occur with increasing age in animals ranging from invertebrates, e.g., yeast and Drosophila, to humans. Carbonylated proteins accumulate over time to form high molecular weight insoluble aggregates if they are escaped degradation by the proteasome. Accumulation of insoluble aggregates has been shown to be extremely toxic to cells and to be associated with several age-related diseases, especially neurodegenerative diseases. Recent work in my laboratory on various short- (lab mouse, wild caught mouse, rat) and long-lived (naked mole rat, bat and marmoset) mammal species including inbred ad libitum and dietary (DR) restricted young and old C57BL/6 mice has shown for the first time an unexpected generalized observation that long- lived species have low level of insoluble cellular protein carbonyl than short-lived species. This data strongly suggests that long lifespan species seems to have better ability to attenuate accumulation of insoluble protein carbonyl than short-lived species. Since carbonylated proteins in general are marked for proteolysis by the proteasome, we speculated that active proteasomal function might be one of the defense machineries used by long-lived species to maintain good quality of proteins. Our preliminary study with rodent species found a direct association between active proteasomal function and increased longevity. Taken these together, the proposed research would expand these intriguing initial observations to evaluate the general hypothesis that long-lived species exhibit reduced accumulation of insoluble protein carbonyl and maintain cell viability via activation of proteasomal machinery. In this proposal, we would like to use variety of non-traditional short- and long-lived species fibroblast cells to test the hypothesis. This project represents the first study to critically evaluate the crosstalk between protein carbonylation and proteasomal function which play a critical role in protecting cells from protein toxicity across a broad range of mammals. These data will allow us to determine if reduced protein toxicity is a common/public mechanism used by evolutionary processes to increase longevity. This proposal will test the hypothesis by pursuing the following specific aims. Specific Aim 1. To test the hypothesis that efficient response of proteasomal function, reduction in insoluble protein carbonyl and restoring cell viability are common traits for fibroblast cells of long-lived species n response to oxidative stress. Specific Aim 2. To test the hypothesis that inactivation of proteasomal function initiates accumulation of insoluble protein carbonyl which lead to increase cell death in fibroblast of long-lived species.