Dissecting the roles of protein O-GlcNAcylation in Toxoplasma gondii

描述（由申请人提供）：刚地弓形虫是顶复门的专性细胞内寄生虫。这种寄生虫在艾滋病患者和免疫功能低下者中引起危及生命的疾病，并在妇女怀孕期间感染时导致出生缺陷或流产。刚地弓形虫在人类宿主中发病和持续的一个重要过程是快速分裂的速殖体和潜伏的慢殖体之间的相互转化。尽管其病理意义，信号分子和这一过程背后的机制知之甚少。最近，在弓形虫中发现了普遍存在的o - glcna酰化和O-GlcNAc转移酶（OGT），这种酶催化了这一过程。OGT在细胞内蛋白质的丝氨酸或苏氨酸残基上添加单个- n -乙酰氨基葡萄糖。由于在哺乳动物生物中，O-GlcNAc是细胞内信号传导、转录调节和响应细胞营养或应激的蛋白质运输的介质，我们假设O-GlcNAc可能通过动态修饰细胞内蛋白质，在弓形虫在快殖子和慢殖子之间的生命周期转换中发挥积极作用。为了验证这一假设，我们将使用化学酶的方法来鉴定弓形虫速殖子中的o - glcnac修饰蛋白（目的1）。接下来，我们将应用该方法对刚地弓形虫速殖子-慢殖子过渡过程中O-GlcNAc蛋白状态的变化进行比较分析。我们将重点研究参与这一过程的基因调控蛋白。一旦确定，我们就会

DESCRIPTION (provided by applicant): Toxoplasma gondii is an obligate intracellular parasite in the phylum Apicomplexa. This parasite causes life-threatening diseases in AIDS patients and immune-compromised individuals, and birth defects or abortions when women are infected during pregnancy. An essential process for pathogenesis and persistence of T. gondii in human hosts is the interconversion between the rapidly dividing tachyzoites and latent bradyzoites. Despite its pathological significance, the signaling molecules and the mechanisms underlying this process are poorly understood. Recently, ubiquitous O-GlcNAcylation and O-GlcNAc transferase (OGT), the enzyme that catalyzes this process, have been identified in T. gondii. OGT adds a single ¿-N-acetylglucosamine to serine or threonine residues of intracellular proteins. Because in mammalian organisms, O-GlcNAc is a mediator of intracellular signaling, transcription regulation, and protein trafficking in response to cellular nutrients or stress, we hypothesize that O-GlcNAc may serve as an active player in T. gondii's life cycle transition between tachyzoites and bradyzoites by dynamically modifying intracellular proteins. To test this hypothesis, we will use a chemoenzymatic approach to identify O-GlcNAc-modified proteins in T. gondii tachyzoites (Aim 1). Next, we will apply this approach for comparative analysis of changes in protein O-GlcNAc status in T. gondii tachyzoite-bradyzoites transition. We will focus our studies on gene-regulatory proteins that are involved in this process. Once identified, we will characterize the role of O-GlcNAc in mediating the biological functions of these modified proteins in this life-cycle transition (Aim 2). Finally, we will characterize how changes in global protein O-GlcNAcylation mediated by OGT influence the life-cycle transition of T. gondii (Aim 3).