Epitranscriptomics in the AIDS-opportunistic pathogen Toxoplasma gondii

弓形虫是一种细胞内寄生虫，感染了多达三分之一的人口。病原体 可能导致先天出生缺陷和危及生命的艾滋病毒/艾滋病机会性感染。复制者 阶段(速殖子)发展为潜伏期(缓殖子)，驻留在大脑、心脏和骨骼肌内。 组织，不受免疫和目前批准的抗寄生虫药物的影响。组织囊肿会引起 免疫功能低下患者的感染反复激活，造成艾滋病毒/艾滋病的慢性病 病人。尽管它在临床上很重要，但我们对其分子细节知之甚少。 速殖子和缓殖子之间的转换。研究表明，缓殖子的发育， 在体外可以由细胞应激触发，这是一个复杂的过程，受到转录调控 转录后水平。最近，其他物种的研究表明，mRNA的修饰， 特别是6位腺苷转录后甲基化(M6A)，通过以下方式影响基因表达 改变RNA的加工或翻译。M6A标记的变化与调节细胞 压力、发展和分化。M6A基因的发现代表了基因调控的新层面。 称为表位转录组学，在原生动物寄生虫中尚未被研究过。在初步研究中，我们 确定弓形虫RNA受M6A的影响，我们已经开始表征这种酶复合体 提供这种RNA修饰。在艾滋病毒/艾滋病的背景下研究弓形虫中的m6A具有重要的意义 这种修饰与压力诱导的细胞变化有关，这使得m6A很有可能 参与缓殖子的开发。在这个新的R21中，我们将解决我们的假设，即m6A mRNA 通过识别M6A的变化，将速殖子分化为缓殖子需要进行修饰 在阶段转换过程中的表位转录组学(目标1)和确定传递 M6A与弓形虫mR-NA结合(目的2)。我们建议的研究将是第一次分析m6A在mRNA上的表达。 寄生虫，并将产生有价值的数据集和试剂，以询问这一至关重要的新领域 与引起慢性弓形虫病的组织包囊发展有关的调查 艾滋病毒/艾滋病患者。

Toxoplasma gondii is an intracellular parasite that has infected up to one-third of the population. The pathogen can cause congenital birth defects and life-threatening opportunistic infection in HIV/AIDS. The replicative stage (tachyzoite) develops into a latent stage (bradyzoite) that resides inside brain, heart, and skeletal muscle tissues, and is impervious to immunity and currently approved antiparasitic drugs. Tissue cysts give rise to recurrent reactivation of infection in immunocompromised patients, creating chronic disease in HIV/AIDS patients. Despite its clinical importance, we know very little about the molecular details orchestrating the conversion between tachyzoites and bradyzoites. Studies have revealed that the development of bradyzoites, which can be triggered in vitro by cellular stress, is a complex process subject to regulation at transcriptional and post-transcriptional levels. Recently, it has been shown in other species that mRNA modifications, specifically post-transcriptional methylation of adenosines at position 6 (m6A), influence gene expression by altering RNA processing or translation. Changes in m6A marks have been associated with modulating cellular stress, development, and differentiation. The discovery of m6A represents a new layer of gene regulation called epitranscriptomics that has not been investigated in protozoan parasites. In preliminary studies, we establish that Toxoplasma RNA is subject to m6A, and we have begun characterizing the enzyme complex that delivers this RNA modification. The study of m6A in Toxoplasma in the context of HIV/AIDS is significant as this modification has been linked to stress-induced changes in cells, making it highly likely that m6A participates in bradyzoite development. In this new R21, we will address our hypothesis that m6A mRNA modifications are required for tachyzoite differentiation into bradyzoites by identifying changes in the m6A epitranscriptome during stage conversion (Aim 1) and determining the “writer” protein complex that delivers m6A onto Toxoplasma mRNA (Aim 2). Our proposed studies will be the first analysis of m6A on mRNA in parasites, and will generate valuable datasets and reagents needed to interrogate this vital new area of investigation relevant to the development of the tissue cysts, which give rise to chronic toxoplasmosis in HIV/AIDS patients.