A Simple Cellular Model for Lipodystrophy

项目总结 胞浆脂滴在细胞器中是独一无二的。这些基本的结构，几乎在所有 真核细胞除了储存脂肪外，还具有多种功能，包括作为荷尔蒙和 信号分子，为其他细胞器提供磷脂前体，天然免疫，并作为平台 用来组装病原体。与其他细胞器不同，液滴包含一个磷脂小叶，该小叶 围绕着它们的疏水核心。尽管事实是它们都是肥胖的基础，这影响了两个人 三分之一的美国人患有脂肪营养不良症，这是一种由于不能正确储存脂肪而导致的致命疾病 对它们的组装仍然是粗略的了解。最近的进展表明，虽然液滴可以形成 一些蛋白质和脂类自发地参与调节这一过程，并确保充分 产生了功能液滴。虽然这些组件已经确定，但它们的许多方面都是未知的 功能。这项建议旨在测试有关Pet10p(一种酵母)作用机制的假设 Perilipin)、Seipin和Scs3p(一种Fit2蛋白)，并从分子上深入了解它们如何共同发挥作用。一个 驱动假说是Pet10p在反式作用中合作(即在蕾期而不是在内质网/液滴连接处) 与Seipin、Scs3p和二酰甘油一起促进和调节液滴组装。在目标1中，重点是 Pet10p，PET10缺失细胞中的多种表型需要与特定的结构域匹配来探测 功能。将阐明Pet10P对液滴稳定作用的基础，以及是否相同 活性对其在液滴形成中的作用将被确定。目标2的重点将是一个更深层次的 了解Seipin的功能，包括将功能映射到结构，靶向蛋白的控制 液滴及其在液滴矢量萌发中的作用。从低温电子显微镜图像中阐明结构的研究将 也是启动的。目标3将重点放在Seipin、Pet10p和Scs3p在液滴的早期步骤上的合作 萌芽中。Seipin是否通过Seipin的氨基末端吸引Pet10p将被确定，以及Seipin的作用 将研究Pet10p和Scs3p对发芽的影响。将探讨到达这个新生结构的顺序。 总体而言，拟议的研究将阐明液滴组装因子的重要功能，并且对于 第一次，研究他们是如何互动的。

PROJECT SUMMARY Cytosolic lipid droplets are unique among cellular organelles. These essential structures, found in nearly all eukaryotic cells, serve several functions besides fat storage, including serving as a depot for hormones and signaling molecules, providing phospholipid precursors for other organelles, innate immunity, and as a platform for assembly of pathogens. Unlike other organelles, droplets contain a single phospholipid leaflet which surrounds their hydrophobic cores. Despite the fact that they are the basis of both obesity, which affects two thirds of Americans, and lipodystrophy, an often fatal disease caused by the inability to properly store fat, there is still sketchy knowledge of their assembly. Recent progress has shown that, while droplets can form spontaneously, a few proteins and lipids are involved in regulating this process and ensuring that fully functional droplets are produced. While these components have been identified, much is unknown about their functions. This proposal seeks to test hypotheses regarding the mechanism of action of Pet10p (a yeast perilipin), seipin, and Scs3p (a Fit2 protein), and to gain molecular insight into how they function together. A driving hypothesis is that Pet10p collaborates in trans (i.e., at the bud rather than at the ER/droplet junction) with seipin, Scs3p, and diacylglycerol to promote and regulate droplet assembly. In Aim 1, focusing on Pet10p, the multiple phenotypes in PET10-deletion cells with be matched with specific domains to probe function. The basis of the stabilizing effect of Pet10p on droplets will be elucidated, and whether the same activity is responsible for its role in droplet formation will be determined. The focus of Aim 2 will be a deeper understanding the function of seipin, including mapping function to structure, control of targeting proteins to droplets, and its role in vectorial budding of droplets. Studies to elucidate structure from cryo-EM images will also be initiated. Aim 3 will focus on the collaboration of seipin, Pet10p and Scs3p on early steps of droplet budding. Whether seipin attracts Pet10p through seipin’s amino terminal will be determined, and the role of Pet10p and Scs3p on budding will be studied. The order of arrival at the nascent structure will be probed. Overall, the proposed research will elucidate important functions of the droplet assembly factors, and, for the first time, study how they interact.