摘要
植物病原细菌产生的harpin类蛋白质可以诱导过敏性细胞死亡(hypersensitive cell death，HCD)，增强植物抗旱性，诱导植物抗病、抗虫，促进植物生长。水稻细条斑病菌HpaG_(Xooc)蛋白质作为harpin蛋白家族的一员，包含两个富含甘氨酸的结构域(glycine-rich motif，GRM)，一个半胱氨酸(cysteine)。GRM是harpin类蛋白共有的特征，而半胱氨酸是其它harpin类蛋白所没有的。分离水稻细条斑病菌HpaG_(Xooc)的不同功能域，我们得到其九种功能片段，分别含有全长中的一段氨基酸序列，如HpaG_(1-94)含有HpaG_(Xooc)的1-94位氨基酸。其中，HpaG_(1-94)、HpaG_(10-42)以及HpaG_(62-138)，在促进作物生长、抗病以及诱导HCD等方面均强于HpaG_(Xooc)。本研究以无活性蛋白(EVP)为对照，研究这三种功能片段与HpaG_(Xooc)相比，对绿茶(山茶科，荼属)田间产量、生化品质方面的改良作用。实验表明，HpaG_(Xooc)，尤其是HpaG_(1-94)不仅可以促进绿茶新芽萌发、嫩叶生长，增加茶园产量，而且可以提高叶片中儿茶酚的含量，改良绿茶的生化品质。HpaG_(Xooc)、HpaG_(1-94)处理绿茶，与EVP相比，茶园产量分别提高39％、55％，烘培后叶片中儿茶酚含量分别提高64％、72％。expansin基因调节植物细胞生长，茶树查耳酮合酶CsCHS基因调控茶树中儿茶酚的生物合成。RT-PCR和RNA gel blotting结果表明，HpaG_(Xooc)和HpaG_(1-94)可以诱导绿茶叶片中expansin基因和CsCHS基因表达；HpaG_(1-94)的诱导表达水平要明显高于HpaG_(Xooc)。综上所述，绿茶作为一种有药用价值的饮料，对栽培和加工过程的无公害有着严格的要求；HpaG_(Xooc)尤其是其功能片段HpaG_(1-94)，能够有效提高绿茶田间产量、改良其生化品质，因此有望在绿茶安全改良方面发挥作用。
Harpin proteins from plant pathogenic bacteria can stimulate hypersensitive cell death(HCD), drought tolerance, defense responses against pathogens and insects in plants, and they can enhance plant growth as well. HpaG_(Xooc), produced by Xanthomonas oryzae pv. oryzae, is a member of harpin group of proteins. The protein contains two copies of the glycine-rich motif(GRM), a characteristic of harpins, and a cysteine, which is absent in other harpins. Recently, we have identified nine functional fragments of HpaG_(Xooc), a harpin protein from Xanthomonas oryzae pv. oryzicola, the pathogen that causes bacterial leaf streak in rice. Fragments HpaG_(1-94), HpaG_(10-42), and HpaG_(62-138), which contain HpaG_(Xooc) regions of amino acid sequence as indicated by the number spans, exceed the parent protein in promoting growth, pathogen defense, and HCD in plants. Here we report improved productivity and biochemical properties of green tea(Camellia sinensis) in response to the fragments tested in comparison with HpaG_(Xooc) and an inactive protein control. Filed tests suggested that the four proteins markedly increased growth and yield of green tea, and increased leaf contents of tea catechols, a group of compounds that have relevance in prevention and treatment of human diseases. In particular, HpaG_(1-94) was more active than HpaG_(Xooc) to expedite growth of juvenile buds and leaves used as green tea material and increase contents of catechols in processed teas. When tea shrubs were treated with HpaG_(Xooc) and HpaG_(1-94) compared to control, green tea yields were over 39% and 55% greater, and leaf catechols were increased by more than 64% and 72%, respectively. The expression of three homologues of expansin genes, which regulate plant cell growth, and the CsCHS gene encoding a tea chalcone synthase, which critically regulates biosynthesis of catechols, were induced in germinal leaves of tea plants following treatment with HpaG_(1-94) or HpaG_(Xooc). Higher levels of gene expression were induced by the application of HpaG_(1-94) than HpaG_(Xooc). Our results suggest that the harpin protein, especially the functional fragment HpaG_(1-94), can be used to effectively increase yield and improve the biochemical property of green tea, a drinking material with medical effects and serious requirement for public harmless farming and processing.
引文
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