Lipid droplets as protein sequestration sites

描述（申请人提供）：脂滴是普遍存在的脂肪储存细胞器，在健康和疾病的脂质代谢中起着核心作用。这些复杂的，动态的细胞器已经牵连到细胞功能远远超出脂质稳态。特别是，它们被提议作为有毒或不稳定蛋白质的临时储存地点，作为大分子复合物的组装平台，或作为细胞内递送蛋白质的载体。尽管脂滴在几种病毒的生命周期中明显发挥着重要作用，并可能促进病毒组装，但它们是否作为内源性蛋白质的瞬时蛋白质隔离位点仍未确定。来自其他细胞区室的一长串蛋白质已被证明以一种受调节的方式重新安置到脂滴上，但这些蛋白质是否在液滴表面发挥新功能或确实受到液滴结合的调节尚不清楚。封存模型的一个关键测试对我们理解基本的细胞和发育过程以及由异常脂肪储存引起的人类疾病（包括脂肪营养不良和肥胖）的管理具有重要意义。这个项目的目标是解决这个问题，特别是一个

DESCRIPTION (provided by applicant): Lipid droplets are ubiquitous fat storage organelles and play central roles in lipid metabolism in both health and disease. These complex, dynamic organelles have been implicated in cellular functions well beyond lipid homeostasis. In particular, they have been proposed to serve as temporary storage sites for toxic or unstable proteins, as assembly platforms for macromolecular complexes, or as vehicles to deliver proteins intracellularly. Although lipid droplets clearly play an important role in the life cycle f several viruses and likely promote viral assembly, whether they act as transient protein sequestration site for endogenous proteins remains unresolved. A long list of proteins from other cellular compartments has been shown to relocate to lipid droplets in a regulated manner, but whether these proteins perform novel functions at the droplet surface or are indeed regulated by droplet association is unclear. A critical test of the sequestration model has important implications for our understanding of basic cellular and developmental processes as well as for the management of human diseases resulting from aberrant fat storage, including lipodystrophies and obesity. The goal of this project is to resolve this issue for one particularly provocative case of proposed protein sequestration, histone storage on lipid droplets in early Drosophila embryos. In this model system, it is possible to combine genetics, biochemistry and live imaging to address this general question. In these embryos, massive amounts of certain histones associate transiently with lipid droplets; this association is mediated by the novel protein Jabba, the putative histone anchor on droplets. In the absence of Jabba, histone levels in embryos are dramatically reduced and - when new histone synthesis is mildly compromised - embryos die with phenotypes indicating lack of sufficient histone supplies. These observations support a model that histones are stored on lipid droplets for later use in development. The goal of this project is to test central predictions of this model. Using a structure-function approach, the domains of Jabba that mediate droplet and histone binding will be identified and the physical state of histones on the droplets will be determined. This information will be employed to generate Jabba mutants lacking specific interactions or mislocalized to other cellular compartments; these mutants will be tested for their ability to rescue high histone levels and normal development. Live imaging and droplet transplantation will address how quickly histones are transferred from lipid droplets to nuclei and whether Jabba remains stably behind. Finally, histone overexpression will be employed to address if droplets normally buffer the histone supply and protect against histone overabundance. If successful, the proposed studies will provide novel insights into both histone metabolism and lipid-droplet function. Most importantly, these studies will provide a paradigm for how and why proteins from other compartments are transiently sequestered on lipid droplets.