CONTROL OF EUCARYOTIC FUNCTION BY METHYLATION

描述(申请人提供)：我们将研究生理功能是如何由甲基化反应控制的，重点是催化蛋白质精氨酸、赖氨酸和异天冬氨酸残基修饰的酶。我们特别感兴趣的是甲基转移酶，它们参与信号转导和代谢调节途径，在这些途径中，甲基的存在或不存在可以调节接受甲基的物种的功能。这些反应对于保护生物体免受环境压力和老化细胞中自发损伤的积累具有重要意义。我们还将开发方法来识别新类型的甲基转移酶，这些甲基转移酶可能在这些途径中催化以前未被识别的反应。我们将确定蛋白质精氨酸甲基转移酶(PRMTs)真核家族的新成员的酶学和功能作用。这些酶改变了精氨酸残基与RNA、DNA和蛋白质伙伴相互作用的能力，并已被证明在基因调节、DNA修复和细胞内信号通路中发挥作用。我们将把我们的工作重点放在哺乳动物系统上，但也会使用酵母和锥虫作为模型系统。我们将特别关注与肿瘤形成和干细胞存活有关的人类PRMT7蛋白。我们的总体目标是建立自然界中催化这些修饰的酶的完整特征铸型，以便充分了解它们的功能，特别是在健康和疾病中的信号和基因调节。我们将表征蛋白修复L-异天冬氨酸/D-天冬氨酸甲基转移酶在细胞内信号转导中的新作用。在这里，我们将利用小鼠和蠕虫(秀丽线虫)系统来探索年龄受损蛋白质的积累、蛋白质修复甲基转移酶对它们的识别以及胰岛素/胰岛素样信号系统对提高抗逆性和寿命的反应之间的关系。我们将测试几个假说来解释该连锁的生理作用，包括通过信号通路本身或上调信号通路的一个或多个关键成员的转录系统直接识别受损蛋白质或甲基化蛋白质。我们将确定赖氨酸蛋白甲基化反应在翻译装置中的作用，包括核糖体蛋白和延伸因子。我们将在酵母和哺乳动物细胞中工作，以了解核糖体蛋白和eEF1A的修饰如何有助于翻译控制和对环境毒素的抗性。最后，我们将使用生物信息学和生化方法来寻找和表征酵母和人类中的新型甲基转移酶。我们特别感兴趣的是确定与人类血浆同型半胱氨酸水平升高相关的甲基转移酶抑制的潜在新部位，这些部位与心血管和神经疾病有关。

DESCRIPTION (provided by applicant): We will investigate how physiological functions are controlled by methylation reactions, focusing on the enzymes that catalyze the modification of protein arginine, lysine, and isoaspartyl residues. We are especially interested in methyltransferases that are involved in signal transduction and metabolic regulatory pathways where the presence or absence of the methyl group can modulate the function of the methyl-accepting species. These reactions are important in protecting organisms from environmental stresses and from the accumulation of spontaneous damage in aging cells. We will also develop methods to identify new types of methyltransferases that may catalyze previously unrecognized reactions in these pathways. We will determine the enzymology and functional roles of new members of the eucaryotic family of protein arginine methyltransferases (PRMTs). These enzymes alter the ability of the arginine residue to interact with RNA, DNA, and protein partners and have been shown to have roles in gene regulation, DNA repair, and intracellular signaling pathways. We will focus our work on mammalian systems, but will also use yeast and trypanosomes as model systems. We will pay special attention to the human PRMT7 protein that has been implicated in tumor formation and stem cell survival. Our overall goal is to establish the complete cast of characters of the enzymes that catalyze these modifications in nature so that their functions, especially in signaling and gene regulation in health and disease, can be fully understood. We will characterize new roles in intracellular signaling for the protein repair L-isoaspartyl/D-aspartyl methyltransferase. Here, we will utilize both mouse and worm (Caenorhabditis elegans) systems to explore the relationships between the accumulation of age-damaged proteins, their recognition by the protein repair methyltransferase, and the responses of the insulin/insulin-like signaling system to increase stress resistance and longevity. We will test several hypotheses to explain the physiological role of the linkage, including direct recognition of damaged proteins or methylated proteins by either the signaling pathway itself or a transcriptional system that upregulates one or more crucial members of the signaling pathway. We will determine the role of lysine protein methylation reactions in the translational apparatus, including ribosomal proteins and elongation factors. We will work in both yeast and mammalian cells to understand how the modifications of ribosomal proteins and eEF1A contribute to translational control and resistance to environmental toxins. Finally, we will use bioinformatic and biochemical approaches to search for and to characterize new types of methyltransferases in both yeast and humans. We are especially interested in identifying potential novel sites of methyltransferase inhibition associated with elevated plasma homocysteine levels in humans that have been linked to cardiovascular and neurological diseases.