Role of host cell metabolism in supporting intracellular Trypanosma cruzi growth

描述(申请人提供)：克氏锥虫是一种引起人类恰加斯病的原生动物寄生虫，是一种专性的细胞内病原体，生活在其哺乳动物宿主细胞的细胞质中，并在细胞质中分裂。尽管它在组织感染、持久性和疾病中很重要，但目前对T ruzi细胞内无鞭毛体阶段的生物学知之甚少。特别是，宿主细胞对支持克氏毛滴虫细胞内复制和生存的基本贡献在很大程度上仍不清楚。为了确定克氏毛滴虫细胞内感染的关键热细胞调节因子，我的实验室建立并进行了高通量全基因组功能筛选，以确定作为细胞内寄生虫生长调节因子的宿主细胞途径/过程。宿主细胞脂肪酸代谢出现在 这是调节HeLa细胞内克氏毛滴虫无鞭毛体生长的主要途径。我们还观察到，细胞内的克氏锥虫无鞭毛体隔离宿主细胞的糖原，其中糖原颗粒在胞质无鞭毛体附近高度浓缩。这些新的发现强烈暗示了克鲁兹毛滴虫在感染的宿主细胞中调节宿主代谢功能的能力。为了支持这一假说，细胞内葡萄糖和脂肪酸利用(有利于脂肪酸利用)之间的主要调节因子PDK4被证明对支持细胞内T.ruzi的生长非常重要。这很耐人寻味，因为克氏锥虫在体内对偏向于利用脂肪酸的肌肉细胞表现出趋向性。虽然已有许多工具用于研究哺乳动物细胞中的新陈代谢，但这些工具尚未被应用于研究克氏毛滴虫感染背景下的宿主细胞新陈代谢。因此，这一探索性的R21提案的目的是建立一个实验框架，其中包括主要为代谢性疾病(糖尿病、癌症等)研究开发的方法和工具，使我们能够确定感染旋毛虫的宿主细胞的宿主细胞代谢发生变化的程度，以及这如何影响细胞内寄生虫的生长。了解寄主代谢和克氏锥虫无鞭毛虫生长之间的密切关系，将提供目前我们对克氏锥虫-宿主相互作用缺乏的重要生物学知识，并将有助于阐明控制克氏锥虫生长的新靶标。 公共卫生相关性：我们的研究重点是克氏锥虫，这是一种导致人类恰加斯病的原虫寄生虫，是全球慢性感染性心肌炎的主要原因。鉴于哺乳动物宿主细胞在支持细胞内毛滴虫生长和生存方面的重要作用，可以预测宿主和寄生虫的代谢过程是密切相关的。我们最近的数据为宿主脂肪酸代谢在支持克氏锥虫细胞内生长中的作用提供了功能证据。这项研究的目的是建立一个实验框架，在这个框架内，我们可以检验关于宿主细胞代谢在支持克氏锥虫生长中的作用的具体假设。

DESCRIPTION (provided by applicant): Trypanosoma cruzi, the protozoan parasite that causes human Chagas' disease, is an obligate intracellular pathogen that lives and divides in the cytoplasm of its mammalian host cell. Despite its importance in tissue infection, persistence and disease, little is currently known about the biology of the intracellular amastigote stage of T cruzi. In particular, the essential contributions of the host cell toward the support of intracelluar replication and survival of T. cruzi remain largely unknown. With a view to identifying critical hot cell regulators of intracellular T. cruzi infection, my laboratory has established and conducted a high-throughput genome-wide functional screen to identify host cell pathways/processes that function as regulators of intracellular parasite growth. Host cell fatty acid metabolism emerged in this screen as a major pathway regulating growth of intracellular T. cruzi amastigotes in HeLa cells. We also made the observation that intracellular T. cruzi amastigotes sequester host cell glycogen, where glycogen particles are highly enriched in the vicinity of the cytosolic amastigotes. These novel findings are strongly suggestive of the ability of T. cruzi to modulate host metabolic functions in infected host cells. In support of this hypothesis, PDK4, the main regulator of the shift between glucose and fatty acid utilization in cells (favoring fatty acid utilization) was shown to be important for supporting intracellular T. cruzi growth. This is intriguing given that T. cruzi exhibit tropism in vivo for muscle cells that are biased toward fatt acid utilization. While many of the tools exist to study metabolism in mammalian cells, these tools have yet to be applied to the study of host cell metabolism in the context of T. cruzi infection. Thus, the aim of this exploratory R21 proposal is to establish an experimental framework with methodologies and tools developed primarily for studies of metabolic disease (diabetes, cancer etc) that will allow us to determine the extent to which host cell metabolism is altered in T. cruzi-infected host cells and how this impacts intracellular parasite growth. Understanding the intimate relationship between host metabolism and T. cruzi amastigote growth will provide a critical piece of biology that is currently lacking in our knowledge of T. cruzi-host interactions and will help to elucidate novel targets for the control of T. cruzi growth PUBLIC HEALTH RELEVANCE: Our studies focus on Trypanosoma cruzi, the protozoan parasite that causes human Chagas' disease, the leading cause of chronic infectious myocarditis worldwide. Given the essential role of the mammalian host cell in supporting growth and survival of intracellular T. cruzi, it is predicted that the host and parasite metabolic processes are intimately linked. Our recent data provide functional evidence for the role of host fatty acid metabolism in supporting intracellular growth of T. cruzi. The aim of the proposed study is to establish the experimental framework within which we can test specific hypotheses regarding the role of host cellular metabolism in supporting T. cruzi growth.