Structure-based biochemical understanding of Sestrins in aging and metabolism

 描述(申请人提供)：衰老是细胞和身体功能逐渐衰退的过程。许多人类疾病，如癌症、II型糖尿病、神经退行性疾病，都与衰老直接或间接相关。因此，老龄化和与年龄相关的疾病会对日常生活质量产生重大影响，尤其是在老年群体中。Sestrin(SESN)是一个应激诱导基因家族，可在多种环境胁迫下上调，包括DNA损伤、氧化应激、低氧和未折叠蛋白质应激。Sesn有两个重要的生物活性，即减少活性氧自由基(ROS)和抑制mTOR复合体1(MTORC1)，这两种活性都可以延缓衰老及其相关的病理过程。事实上，在包括蠕虫、苍蝇和小鼠在内的许多模型动物中，Sestrin家族蛋白被证明是代谢稳态的关键调节因子，可以减弱与年龄和肥胖相关的各种病理。这些结果表明，七叶皂苷是进化上保守的抗衰老分子。然而，由于Sestrins的这些抗衰老活性的生化基础一直难以捉摸，我们无法利用Sestrins的有益活性来延缓衰老和延长健康寿命。基于我们最近通过X射线结晶学测定的人Sestrin2的三维分子结构，我们首次提出了揭示Sestrin蛋白抗衰老活性的生化机制。在目标1中，我们将通过结构导向突变和随后的体内外氧化还原活性的测定来揭示hSesn2‘S抗氧化功能的生化基础。在目标2中，使用从目标1研究中产生的突变的hSesn2蛋白，我们将检查突变的残基和活性位点是否对在培养的哺乳动物细胞和完整动物(果蝇)的组织中抑制mTORC1重要。最后，在目标3中，表现出不同加速衰老表型的dSesn缺失突变果蝇将与野生型和突变型hSesn2重组，以测试突变的关键残基(S)是否在抗衰老生理作用中具有重要功能 塞森。这项研究的成功完成将使我们能够阐明hSesn2在抑制衰老和控制代谢方面的生理功能的结构基础。从这项研究中获得的结构和机理信息不仅揭示了Sesns的抗氧化和mTOR抑制活性的机制，而且还使新的小分子的开发成为可能，这些小分子可以增强Sesns的催化活性或提高Sesn的稳定性。这些分子将在未来开发，可能被用来在药理上延长健康寿命，提高晚年的生活质量。

 DESCRIPTION (provided by applicant): Aging is a process of gradual decline in cellular and bodily function. Many human diseases, such as cancers, type II diabetes, neurodegenerative diseases, are either directly or indirectly associated with aging. Therefore, aging and age-associated disease can significantly influence on the quality of daily life especially within the elderly age group. Sestrin (Sesn) is a stress-inducible gene family that can be upregulated by a variety of environmental stresses including DNA damage, oxidative stresses, hypoxia and unfolded protein stresses. Sesn has two important biological activities in reducing reactive oxygen species (ROS) and suppressing mTOR complex 1 (mTORC1), both of which may attenuate aging and its associated pathologies. Indeed, in many model animals, including worms, flies and mice, Sestrin-family proteins were shown to be a critical regulator of metabolic homeostasis that attenuates diverse age- and obesity-associated pathologies. These results suggest Sestrins to be evolutionarily conserved anti-aging molecules. However, because the biochemical basis for these anti-aging activities of Sestrins has been elusive, we were unable to harness Sestrins' beneficial activities for attenuation of aging and extension of healthspan. Based on the 3D molecular structure of human Sestrin2, which we have recently determined through X-ray crystallography, here we propose to uncover the biochemical mechanisms underlying the anti-aging activity of Sestrin proteins for the first time. In Aim 1, we will revealthe biochemical basis underlying hSesn2's antioxidant function using structure-guided mutagenesis and subsequent in vitro and in vivo assays of its redox activity. In Aim 2, using the mutant hSesn2 proteins generated from Aim 1 research, we will examine whether the mutated residues and active sites are important for suppressing mTORC1 in cultured mammalian cells and in tissues of an intact animal (Drosophila). Finally in Aim 3, dSesn-null mutant flies, which exhibit diverse accelerated aging phenotypes, will be reconstituted with wild-type and mutant hSesn2 to test if the mutated key residue(s) are functionally important for anti-aging physiological roles of Sesn. Successful completion of the proposed research will allow us to elucidate the structural basis for the physiological function of hSesn2 in suppressing aging and controlling metabolism. The structural and mechanistic information obtained from the proposed study not only reveals the mechanisms underlying Sesns' antioxidant and mTOR-inhibiting activities, but also enables development of novel small molecules that can either enhance the catalytic activity of Sesns or increase Sesn stability. These molecules, which will be developed in future, may be used to pharmacologically expand healthspan and improve the quality-of-life in the later ages.