Vacuolar Protein Sorting in Plant Cells

植物细胞中的液泡蛋白质分选

• 批准号: 9315940
• 负责人: Maarten Chrispeels
• 金额: $ 39.33万
• 依托单位: University of California-San Diego
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 1994
• 资助国家: 美国
• 起止时间: 1994-02-01 至 1998-01-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=9315940&HistoricalAwards=false
• 关键词: Vacuolar; Protein; Sorting; Plant; Cells

9315940 Chrispeels, Marten J. The goal of the proposal is to understand how proteins are targeted to and accumulated in vacuoles of plant cells. Although plant vacuoles are related to yeast vacuoles and lysosomes of mammalian cells, they contain a much greater variety of proteins, including storage proteins, lectins, enzyme inhibitors, defense enzymes, as well as many of the acidic hydrolases commonly found in all vacuoles and lysosomes. Much progress has been made in the last years in understanding the sorting signals that occur on the vacuolar proteins themselves, but there is at present no information on the cellular machinery that allows these proteins to be correctly targeted to their destination. Correct vacuolar targeting of Golgi-derived vesicles involves proteins on the outside of the vesicles that mediate vesicle budding, transport and fusion of the vesicles with the target membrane in a highly specific manner. These processes are thought to require coating and uncoating with proteins that bind to the membranes and can be recruited from the cytosol. The identification of these proteins is the major goal of this proposal. The Pi will exploit the recent isolation of the gene that encodes a phosphatidyl-inositol 3-kinase from Arabidopsis and investigate whether it has a role in vacuolar protein sorting. The function of this protein will be examined in stably transformed and transient systems, or protoplasts treated with antisense oligodeoxyribonucleotides. The Pi will transform a yeast vps34 mutant with the plant phosphatidylinositol 3-kinase gene to find interacting proteins, and then rescue this strain with a plant cDNA library. The two-hybrid method is also going to be used to identify this gene. The Pi will also continue work on the targeting of proteins to the vacuolar membrane or tonoplast. He will make constructs that involve a reporter protein against which he has antiserum, and different membrane spanning domains. %%% The significance of this work lies in a better understanding of vacuole biogenesis in plant cells. The research proposed here will contribute to our understanding of the mechanisms by which the vacuolar proteins reach their destinations. As with other aspects of the secretory system, some of the findings will be applicable to all eukaryotic cells, while others will be specific for plant cells. ***

9315940 Marten J.克里斯皮尔斯该提案的目标是了解蛋白质是如何被定位到植物细胞的液泡中并在其中积累的。虽然植物液泡与哺乳动物细胞中的酵母液泡和溶酶体有关，但它们含有种类更多的蛋白质，包括贮藏蛋白、凝集素、酶抑制剂、防御酶以及在所有液泡和溶酶体中普遍存在的许多酸性水解酶。在过去的几年里，在理解发生在液泡蛋白上的分选信号方面已经取得了很大的进展，但目前还没有关于细胞机制的信息，使得这些蛋白质能够正确地靶向它们的目的地。高尔基体小泡的正确空泡靶向涉及小泡外部的蛋白质，这些蛋白质以高度特异的方式介导小泡的萌发、运输和小泡与靶膜的融合。这些过程被认为需要用结合到膜上的蛋白质包覆和去掉包被，这些蛋白质可以从细胞质中招募。鉴定这些蛋白质是这项提议的主要目标。PI将利用最近从拟南芥中分离到的编码磷脂酰肌醇3-激酶的基因，并研究它是否在空泡蛋白分类中发挥作用。这种蛋白质的功能将在稳定转化和瞬时系统中进行检测，或者用反义寡核苷酸处理原生质体。PI将转化带有植物磷脂酰肌醇3-激酶基因的酵母Vps34突变体，以寻找相互作用的蛋白质，然后用植物cDNA文库拯救该菌株。双杂交方法也将被用于鉴定该基因。PI还将继续将蛋白质靶向液泡膜或液泡膜的工作。他将构建一种结构，涉及一种他有抗血清的报告蛋白，以及不同的跨膜结构域。这项工作的意义在于更好地理解植物细胞中液泡的生物发生。这里提出的研究将有助于我们理解液泡蛋白到达目的地的机制。与分泌系统的其他方面一样，一些发现将适用于所有真核细胞，而另一些将专门针对植物细胞。***