FUCTIONAL DIFFERENTIATION OF PLANT VACUOLES

植物液泡的功能分化

基本信息

批准号：
10440244
负责人：
NISHIMURA Ikuko hara
金额：
$ 8.51万
依托单位：
NATIONAL INSTITUTE FOR BASIC BIOLOGY
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
1998
资助国家：
日本
起止时间：
1998 至 1999
项目状态：
已结题

项目摘要

Novel vesicles that accumulate large amounts of proprotein precursors of storage proteins were purified from maturing pumpkin seeds. These vesicles were designated precursor-accumulating (PAC) vesicles and *ave diameters of 200 to 400 nm. They contain an electron-dense core of storage proteins surrounded by an electron-translucent layer, and some vesicles also contained small vesicle-like structures. Numerous electron-dense aggregates of storage proteins within the endoplasmic reticulum were found to be develop into the electron-dense cores of the PAC vesicles and then leave the endoplasmic reticulum. The unique PAC vesicles might mediate a transport pathway for insoluble aggregates of storage proteins directly to protein storage vacuoles. We also found that PAC vesicle can be induced in vegetative cells by ectopic expression of the protein that is destined to be compartmentalized into the PAC vesicles.Inactive precursors that are accumulated in the PAC vesicles should be converted int … More o mature forms. Vacuolar processing enzyme (VPE) has been shown to be responsible for maturation of various seed proteins in plant vacuoles. Arabidopsis has three VPE homologues; βVPE is specific to seeds and αVPE and γVPE are specific to vegetative organs. We found that the vegetative VPE has a vacuolar processing activity and is localized in the lytic vacuoles. The mRNA levels of both vegetative VPEs were increased in the primary leaves during senescence in parallel with the increase of the mRNA level of a senescence- associated gene (SAG2). The vegetative VPE might regulate the activation of some functional vacuolar proteins that are known to respond to these treatments.To investigate a VPE system in protein-storage vacuoles, we isolated the PAC vesicles and characterized a l00-kDa component (PVIOO) of the vesicles. PVIOO was composed of a hydrophobic signal peptide and the following three domains: an 11-kDa Cysrich domain with four CxxxC motifs (C, Cys), a 34-kDa Arg/Glu-rich domain composed of six homologous repeats, and a 50-kDa vicilin-like domain. Two Cysrich peptides, three Arg/Glu-rich peptides and the vicilin-like protein were produced by cleaving Asn-Gin bonds of PV100 and that all these proteins had a pyroglutamate at their NH2 terminus. VPE was responsible for cleaving Asn-Gin bonds of a single precursor, PVIOO, to produce multiple seed proteins. It is likely that the Asn-Gin stretches not only provide cleavage sites for VPE but also produce aminopeptidase-resistant proteins. Cys-rich peptide function as a trypsin inhibitor and Arg/Glu-rich peptides function as cytotoxic peptides. Our findings suggested that PVIOO is converted into different functional proteins in the vacuoles of seed cells. Less

从成熟的南瓜种子中纯化了积累大量储存蛋白的大量提谋蛋白前体的新型蔬菜。这些蔬菜的设计前体蓄积（PAC）蔬菜和 *AVE直径为200至400 nm。它们包含一个被电子透明层包围的储存蛋白的电子密集核心，一些蔬菜还包含小蔬菜样结构。发现内质网中储存蛋白的许多电子致密聚集体被发现发展为PAC蔬菜的电子致密核心，然后离开内质网。独特的PAC蔬菜可能会介导一条传输途径，以直接将储存蛋白的不溶性聚集到蛋白质储存真空中。我们还发现，可以通过注定要分裂为PAC蔬菜的蛋白质的蛋白质的依托蛋白表达PAC蔬菜。液泡加工酶（VPE）已被证明是造成植物真空中各种种子蛋白的成熟的原因。拟南芥具有三个VPE同源物； βVPE特定于种子，αVPE和γVPE特定于营养器官。我们发现营养VPE具有液泡加工活性，并位于裂解液泡中。在感应期间，两种营养VPE的mRNA水平与感应相关基因的mRNA水平的增加（SAG2）同时增加。营养VPE可能会调节某些已知对这些处理的功能性真空蛋白的激活。为了研究蛋白质储存真空糖中的VPE系统，我们分离了PAC蔬菜并表征了蔬菜的L00-KDA成分（PVIOO）。 PVIOO由疏水信号肽和以下三个结构域组成：一个11 kDa Cysrich结构域，具有四个CXXXC基序（C，Cys），一个34 kDa arg/glu-rich域组成，由6个同源重复序列和一个50 kda vicicilin like domain组成。两种Cysrich Petides，三个Arg/Glu富含petides和Vicilin样蛋白是通过裂解PV100的Asn-Gin键产生的，并且所有这些蛋白质在其NH2末端都有焦谷氨酸。 VPE负责分裂单个前体PVIOO的ASN轴键，以产生多种种子蛋白。 ASN-gin伸展可能不仅为VPE提供切割位点，而且还会产生抗肽酶耐药蛋白。富含Cys的肽充当胰蛋白酶抑制剂，富含ARG/GLU的肽充当细胞毒性肽。我们的发现表明，PVIOO在种子细胞真空中转化为不同的功能蛋白。较少的