Molecular mechanism of vacuolar processing system in differentiation of plant cell

液泡功能的动态变化是高等植物细胞分化灵活性和细胞死亡事件的重要原因。本研究的目的是阐明液泡处理酶在细胞程序性死亡和细胞分化过程中负责液泡蛋白成熟和激活的分子机制。(1)获得了两种营养类型的液泡处理酶(αvpe和γvpe)和一种种子型vpe(βvpe)的基因敲除突变体。然后将它们杂交，获得双重和三重突变体。通过对三个突变体的分析发现，正常加工的贮藏蛋白中没有检测到VPEs，说明VPEs是种子贮藏蛋白加工中的关键酶。(2)我们发现一个特征结构来源于ER，ER小体。内质网小体在幼苗的表皮细胞中发育，含有VPE…等应激诱导酶前体蛋白。更多。相反，莲座叶没有内质网小体。我们发现，创伤胁迫诱导了莲座状叶片中大量内质网小体的形成。内质网小体的生物学功能可能与高等植物的防御系统有关。(3)半胱氨酸蛋白酶是控制动物细胞死亡的重要酶。在植物中，许多研究表明，caspase样活性在死亡的细胞中被诱导。为了确定caspase在植物中的功能同源物，我们研究了烟草花叶病毒(TMV)侵染烟草植株的过敏性细胞死亡。VPE特异性抑制剂和caspase-1特异性抑制剂均可阻断TMV诱导的细胞过敏性死亡。由于VPE对caspase-1特异性底物具有caspase活性，因此导致caspase-1样活性的蛋白水解酶被鉴定为VPE。从VPE活性在烟草花叶病毒(TMV)诱导的过敏性反应的早期阶段的出现来看，表明VPE调控了TMV诱导的过敏性细胞死亡的早期过程。较少

Dynamic transformation of vacuolar function is responsible for flexibility of differentiation and cell death events in higher plant cells. The aim of this study is to elucidate the molecular mechanism of vacuolar processing enzyme (VPE), which is known to be responsible for maturation and activation of vacuolar proteins, in the process of programmed cell death and cell differentiation.(1) We generated the Arabidopsis knockout mutants that had a defect of each gene for the two vegetative type VPEs, αVPE and γVPE, and one seed type VPE, βVPE. Then they were crossed to obtain double and triple mutants. From the analysis of the triple mutant, we found that normally processed storage proteins were no more detected, indicating that VPEs are a key enzyme in processing of seed storage proteins.(2) We found that a characteristic structure derived from ER, ER body. ER bodies develop in the epidermal cells of young seeding and contain the precursor proteins of stress-inducible protease such a VPE … More . In contrast, rosette leaves had no ER bodies. We found that wound stress induced the formation of many ER bodies in rosette leaves. It is possible that the biological function of ER bodies is related to defense systems in higher plants.(3) The process of controlled cell death in animals is well organized by caspases that are cysteine proteinases. In plants, many studies have shown that caspase-like activities are induced in dying cells. To identify a functional homolog of caspase in plants, we investigated hypersensitive cell death in tobacco mosaic virus (TMV)-infected tobacco plants. A VPE-specific inhibitor abolished the TMV-induced hypersensitive cell death, as did a caspase-1-specific inhibitor.Because VPE had caspase activity toward a caspase-1-specific substrate, the proteinase responsible for the caspase-1-like activity was identified as VPE. From the appearance of VPE activity at the early stage-of the TMV-induced hypersensitive response in tobacco leaves, it was suggested that VPE regulates the early process of the TMV-induced hypersensitive cell death. Less