Lipid droplets as protein sequestration sites

脂滴作为蛋白质隔离位点

• 批准号: 8642196
• 负责人: MICHAEL Andreas WELTE
• 金额: $ 29.01万
• 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 2017-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8642196
• 关键词: Address; Affect; Anabolism; Atherosclerosis; Binding; Biochemical; Biochemistry; Biological; Biological Models; Biological Testing; Buffers; Cardiovascular Diseases; Caveolins; Cell Nucleus; Cell membrane; Cell physiology; Cessation of life; Characteristics; Complex; Core Protein; DNA Damage; Data; Development; Developmental Process; Diabetes Mellitus; Disease; Drosophila genus; Embryo; Embryonic Development; Ensure; Enzymes; Equilibrium; Fatty acid glycerol esters; Genetic; Goals; Health; Hepatitis; Histones; Homeostasis; Image; Lead; Life; Life Cycle Stages; Link; Lipids; Lipodystrophy; Liver; Macromolecular Complexes; Mammals; Mediating; Metabolism; Modeling; Mutation; Nuclear Proteins; Obesity; Organelles; Phenotype; Play; Production; Proteins; Publishing; Role; Severity of illness; Site; Structure; Surface; Testing; Time; Transplantation; Viral; Viral Proteins; Virus; Wasting Syndrome; Yeasts; design; fly; human disease; in vivo; insight; lipid metabolism; mutant; novel; overexpression; photoactivation; physical state; premature; public health relevance; research study; response

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.

描述（由申请人提供）：脂质液滴是无处不在的脂肪储存细胞器，在健康和疾病中扮演脂质代谢中的核心作用。这些复杂的动态细胞器与脂质稳态以外的细胞功能有关。特别是，已提议它们充当有毒或不稳定蛋白质的临时存储位点，作为大分子复合物的组装平台，或者用作细胞内输送蛋白质的车辆。尽管脂质液滴在生命周期F中显然起着重要作用，并且可能促进病毒组装，但它们是否充当内源性蛋白质的瞬时蛋白质固相位位点仍未解决。已经显示出一长串来自其他细胞室的蛋白质以调节的方式迁移到脂质液滴，但是这些蛋白质是在液滴表面执行新功能还是确实受液滴缔合的调节尚不清楚。对隔离模型的批判性测试对我们对基本细胞和发育过程的理解以及对异常脂肪储存（包括脂肪营养不良和肥胖）产生的人类疾病的管理具有重要意义。该项目的目的是解决这个问题，特别是 提出的蛋白质固相的挑衅性情况，在早期果蝇胚胎中脂质液滴上存储组蛋白。在此模型系统中，可以将遗传学，生物化学和实时成像结合起来，以解决这个普遍的问题。在这些胚胎中，大量的某些组蛋白与脂质液滴瞬时相关。这种关联是由新型蛋白JABBA介导的，Jabba是液滴上的假定组蛋白锚。在没有JABBA的情况下，胚胎中的组蛋白水平大大降低，当新的组蛋白合成受到轻度损害时 - 胚胎死于表型，表明缺乏足够的组蛋白供应。这些观察结果支持一个模型，该模型将组蛋白存储在脂质液滴上，以便以后在开发中使用。该项目的目的是测试该模型的中心预测。使用结构功能方法，将确定介导液滴和组蛋白结合的JABBA的结构域，并确定液滴上组蛋白的物理状态。这些信息将用于产生缺乏特定相互作用或错误定位到其他细胞室的Jabba突变体；这些突变体的能力将被测试，以挽救高组蛋白水平和正常发育。实时成像和液滴移植将解决组蛋白从脂质液滴转移到核的速度，以及JABBA是否保持稳定。最后，将使用组蛋白过表达来解决液滴通常缓冲组蛋白供应并预防组蛋白过度的液滴。如果成功，则提出的研究将提供对组蛋白代谢和脂质滴滴功能的新见解。最重要的是，这些研究将提供一个范式，说明来自其他隔室的蛋白质如何以及为什么在脂质液滴上瞬时隔离。