Roles of endosomal phosphoinositides in intracellular trafficking

内毒素转运是一个受到严格控制的过程，涉及膜融合和裂变事件，这确保了内小体、溶酶体和其他细胞内细胞器或囊泡内货物的正确传递和交换。它对一些细胞功能是必不可少的，如蛋白质和脂类的运输、自噬和溶酶体中大分子的降解。它是如何被调控的是细胞生物学中的一个基本问题。我们的研究计划打算使用病毒作为手段来监测和了解细胞内的囊泡运输。事实上，病毒已经进化出利用甚至调节内体成熟或内溶体运输的机制，因为它可以决定感染是否成功。因此，它们可以成为研究细胞运输途径的强大而宝贵的工具。我们发现，Vac14/Fig4/PIKfyve复合体的蛋白是埃博拉病毒(EBOV)和严重急性呼吸综合征冠状病毒(SARS-CoV)感染所必需的，它调节着内溶体膜上磷脂酰肌醇(3，5)二磷酸的合成。合成PI(3，5)P2的酶复合体是如何被激活的，以及PI(3，5)P2在内酶体转运中的作用尚不清楚。在这里，我们建议使用EBOV和SARS-CoV来回答这些问题。 这项研究计划的目标是了解这种复合体及其产生的肌醇磷脂是如何调节的，以及它们如何控制细胞内溶酶体的运输，这是一个基本的细胞过程。 假设： 内体磷脂酰肌醇PI(3，5)P2对于病毒颗粒运输到发生病毒融合的细胞内间隔是至关重要的。 短期目标： 1)检验遗传和化学操作对PI(3，5)P2水平的影响 2)对Vac14/Fig4/PIKfyve复合体进行分子解剖 3)识别PI(3，5)P2效应器 我们的长期目标是从分子上了解Vac14/Fig4/PIKfyve相互作用组和功能。 为了实现我们的目标，我们将使用遗传学、化学生物学和病毒学工具的组合。将这些特定的工具与我们在分子/细胞生物学和病毒进入方面的专业知识相结合，我们希望填补对内溶酶体贩运以及这一基本过程如何被病毒病原体劫持的理解方面的空白。

Endolysosomal trafficking is a tightly regulated process that involves membrane fusion and fission events, which ensure proper delivery and exchange of cargoes within endosomes, lysosomes, and other intracellular organelles or vesicles. It is essential for several cellular functions such as protein and lipid transport, autophagy, and degradation of macromolecules in lysosomes. How it is regulated is a fundamental issue in cellular biology. Our research program intends to use viruses as means to monitor and understand vesicular trafficking in cells. Indeed, viruses have evolved mechanisms to use and even regulate endosomal maturation or endolysosomal trafficking, since it can determine whether the infection is successful or not. Therefore, they can represent powerful and invaluable tools for the study of cellular trafficking pathways. We found that proteins of the Vac14/Fig4/PIKfyve complex, which regulates the synthesis of the phosphoinositide(3,5)bisphosphate at the endolysosomal membrane, are essential for infection by Ebola virus (EboV) and severe acute respiratory syndrome coronavirus (SARS-CoV). How the enzymatic complex synthesizing PI(3,5)P2 is activated and the role of PI(3,5)P2 in endolysosomal trafficking remain unclear. Here, we propose to use EboV and SARS-CoV to answer these questions. The goal of this research program is to understand how this complex and its resulting phosphoinositide are regulated and how they control endolysosomal trafficking in cells, a fundamental cellular process. Hypothesis: The endosomal phosphoinositide PI(3,5)P2 is critical for the trafficking of viral particles to intracellular compartments where viral fusion occurs. Short-term objectives: 1) Examine the effect of genetic and chemical manipulation of PI(3,5)P2 levels 2) Perform a molecular dissection of the Vac14/Fig4/PIKfyve complex 3) Identify PI(3,5)P2 effectors Our long term goal is to gain a molecular understanding of the Vac14/Fig4/PIKfyve interactome and function. To meet our objectives, we will use a combination of genetic, chemical biology, and virological tools. Combining these specific tools with our expertise in molecular/cell biology and viral entry, we hope to fill a gap in the understanding of endolysosomal trafficking and how this fundamental process can be hijacked by viral pathogens.