Biochemistry, biology and diversity of Fic domains

项目摘要 多种细胞过程通常被破坏以促进癌细胞的增殖， 其中之一是在内质网（ER）中发生的未折叠蛋白反应（UPR）。 重要的是，UPR与炎症或神经元健康之间也有很强的联系。 许多神经退行性疾病和衰老疾病都与普遍定期审议有关。我们有 最近发现了一种新形式的BiP调节，由蛋白质Fic进行的AMP化。我们观察到， 在BiP的ATP结合位点附近的苏氨酸上添加一个腺苷一磷酸（AMP）分子 在正常的生长条件下。这种修饰被相同的酶Fic快速去除， 多种ER应激诱导条件。我们最近已经表明，Fic对BiP的调节是 对维持神经元稳态至关重要。自从我们发现Fic结构域介导 已经鉴定了由细菌Fic结构域进行的AMP化、其他不同的活性。这些 研究已经揭示了Fic结构域在其利用不同底物的能力方面的分子可塑性。 尽管进行了这些研究，但仍有许多不同的Fic蛋白有待表征，无论是在细胞内还是在细胞外。 催化活性、生物学功能和分子靶标。我们提出了三个项目，将进一步 我们对Fic酶的生物学和它们在体外和体内使用的化学的理解。 首先，当ER受到压力时，Fic从AMPylator变为deAMPylator，而关键是 监管正在打破Fic活跃位点的盐桥。我们想了解生物化学 调节这个开关。第二，我们对果蝇遗传学的研究表明，Fic是必需的， 神经可塑性因此，我们有兴趣了解Fic在哺乳动物中起什么作用。 使用转基因小鼠进行生物学研究。第三，有许多不同的Fic蛋白， 在催化活性、生物学功能和分子靶点方面都进行了表征。我们计划 研究这些孤儿Fic结构域并鉴定其生物活性和底物。

Project Summary A variety of cellular processes are commonly subverted to encourage the proliferation of cancer cells, one of which is the unfolded protein response (UPR) that occurs in the endoplasmic reticulum (ER). Importantly, there is also a strong connection between UPR and inflammation or neuronal health. Many neurodegenerative diseases and diseases of aging have connections to the UPR. We have recently discovered a new form of BiP regulation, AMPylation by the protein Fic. We observe that Fic adds an adenosine monophosphate (AMP) molecule to a threonine near the ATP binding site of BiP during normal growth conditions. This modification rapidly is removed by the same enzyme Fic under multiple ER stress-inducing conditions. We recently have shown that the regulation of BiP by Fic is essential for maintaining neuronal homeostasis. Since our discovery of Fic domains that mediate AMPylation, other diverse activities performed by bacterial Fic domains have been identified. These studies have revealed the molecular plasticity of Fic domains in its ability to utilize diverse substrates. Despite these studies, there are many different Fic proteins that remain to be characterized, both in catalytic activity, biological function, and molecular targets. We propose three projects that will further our understanding of the biology of Fic enzymes and the chemistry they use, both in vitro and in vivo. First, when the ER is stressed, Fic changes from an AMPylator to a deAMPylator, and the key to this regulation is breaking a salt bridge in Fic's active site. We want to understand the biochemistry regulating this switch. Second, our studies with Drosophila genetics show that Fic is required for neuronal plasticity. We are therefore interested in understanding what role Fic plays in mammalian biology using transgenic mice. Third, there are many different Fic proteins that remain to be characterized, both in catalytic activity, biological function, and molecular targets. We plan to investigate these orphan Fic domains and identify their biological activities and substrates.