Lipid droplets as protein sequestration sites

脂滴作为蛋白质隔离位点

• 批准号: 9381218
• 负责人: MICHAEL Andreas WELTE
• 金额: $ 38.62万
• 依托单位: UNIVERSITY OF ROCHESTER
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-04-01 至 2021-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9381218
• 关键词: Address; Anabolism; Atherosclerosis; Binding; Biochemical; Biochemistry; Biological; Buffers; Cell Compartmentation; Cell Nucleus; Cell physiology; Cells; Cellular biology; Chromatin Modeling; Cytoplasm; Development; Developmental Process; Diabetes Mellitus; Disease; Dissection; Drosophila genus; Embryo; Embryonic Development; Ensure; Fatty acid glycerol esters; Genetic; Goals; Health; Histones; Homeostasis; Image; Individual; Investigation; Lipids; Lipodystrophy; Modeling; Modification; Molecular; Molecular Genetics; Nuclear; Nurses; Obesity; Oogenesis; Organelles; Organism; Phenotype; Physiological; Play; Post-Translational Protein Processing; Process; Property; Proteins; Proteomics; Regulation; Research; Role; Site; Structure; Surface; System; Testing; Time; Variant; Virus; Work; blastocyst; experimental study; fitness; human disease; imaging approach; insight; lipid metabolism; loss of function; metabolic abnormality assessment; mutant; novel; photoactivation; protein function; proteostasis; tool

Project summary Lipid droplets are intracellular fat storage organelles that play critical roles for lipid and energy homeostasis in both health and disease. Lipid droplets also have a second major function as regulators of protein homeostasis: droplets store specific proteins from other cellular compartments, participate in protein refolding and turnover, and serve as assembly platform for viruses. While there has been great progress in unraveling how droplets control lipid metabolism, studies of mechanisms and physiological relevance of their protein- handling roles remain limited. Yet a better understanding of these processes will not only illuminate many fundamental cellular processes, but may also inform treatment of the myriad of human diseases characterized by over- or understorage of fat. The goal of this project is to mechanistically dissect two proposed general functions of protein sequestration by lipid droplets: long-term storage to safely preserve excess levels of otherwise harmful or unstable proteins and short-term buffering to equalize the availability of proteins despite fluctuations in biosynthesis or demand. This application focuses on an extensively validated example of protein sequestration: histone storage on lipid droplets in early Drosophila embryos. Three key features make this a promising model: First, a crucial component of the sequestration machinery has been defined molecularly, the histone anchor Jabba. Second, buffering activity undergoes a dramatic developmental transition, providing an opportunity to probe the regulation and biological significance of buffering. Third, it is possible to combine classical and molecular genetics, biochemistry, proteomics, and live imaging to study sequestration in an intact, developing organism. Previous work has identified histone- and lipid-droplet binding activities in Jabba. This information will now be employed to determine how these activities promote histone storage, in particular whether Jabba protects histones from degradation or promotes their efficient delivery. Characterization of Jabba variants and photoactivation experiments will determine whether Jabba acts solely as static histone anchor or can accompany histones in the cytoplasm. Using proteomics, post-translational modifications and binding partners of Jabba (and possibly other components) will be determined before and after the transition, with the goal of manipulating the transition and testing the consequences when buffering is turned on or off at inappropriate times. Finally, Jabba is known to promote dispersal of lipid droplets throughout the embryo. The mechanistic basis for this role will be determined, using structure-function approaches and analysis of a putative Jabba binding partner. These investigations will make it possible to test if droplet dispersal enhances buffering or serves other functions, such as modulating lipid metabolism. If successful, these studies will provide paradigms for how lipid droplets can employ regulated protein sequestration to control processes in other compartments of the cell.

项目摘要 脂滴是细胞内的脂肪储存细胞器，其在体内的脂质和能量稳态中起关键作用。 健康和疾病。脂滴还具有第二个主要功能，作为蛋白质的调节剂 稳态：液滴储存来自其他细胞区室的特定蛋白质，参与蛋白质重折叠 和周转，并作为病毒的组装平台。虽然在解开 液滴如何控制脂质代谢，其蛋白质的机制和生理相关性的研究- 处理角色仍然有限。然而，更好地理解这些过程不仅会阐明许多 基本的细胞过程，但也可能告知治疗的无数人类疾病的特点 脂肪储存过多或不足。这个项目的目标是机械地剖析两个建议的一般 脂滴隔离蛋白质的功能：长期储存，以安全地保存过量的 否则有害或不稳定的蛋白质和短期缓冲，以平衡蛋白质的可用性， 生物合成或需求的波动。这个应用程序的重点是一个广泛验证的蛋白质的例子 隔离：果蝇早期胚胎中脂滴上的组蛋白储存。三个关键特征使其成为 有前途的模型：首先，螯合机制的一个关键组成部分已被分子定义， 组蛋白锚贾巴。其次，缓冲活动经历了戏剧性的发展转变，提供了一个 为探讨缓冲作用的调控和生物学意义提供了机会。第三，可以将联合收割机 经典和分子遗传学，生物化学，蛋白质组学和实时成像，以研究隔离在一个 完整的发育中的有机体以前的工作已经确定了贾巴的组蛋白和脂滴结合活动。 这些信息现在将被用来确定这些活动如何促进组蛋白储存，特别是 贾巴是保护组蛋白不被降解，还是促进其高效传递。表征 贾巴变体和光活化实验将确定贾巴是否仅作为静态组蛋白 在细胞质中锚或伴随组蛋白。利用蛋白质组学、翻译后修饰和 Jabba的绑定伙伴（以及可能的其他组件）将在过渡之前和之后确定， 其目的是操纵转换并测试当缓冲被打开或关闭时的结果 不合时宜的时候最后，贾巴被认为可以促进脂滴在整个胚胎中的扩散。的 这一作用的机制基础将被确定，使用结构功能方法和分析的一个 假定的贾巴结合伴侣这些研究将有可能测试液滴扩散是否增强了 缓冲或提供其他功能，如调节脂质代谢。如果成功，这些研究将 为脂滴如何利用受调节的蛋白质螯合来控制 细胞的其他部分。