Tortiglione Claudia, Fogliano Vincenzo, Ferracane Rosalia, Fanti Paolo, Pennacchio Francesco, Monti Luigi Maria, Rao Rosa
Department of Soil, Plant and Environment, University of Naples, via Università 100, Portici 80055, Italy.
Plant Mol Biol. 2003 Dec;53(6):891-902. doi: 10.1023/B:PLAN.0000023667.62501.ef.
Tomato systemin is a signalling peptide produced in response to wounding that locally and systemically activates several defence genes. The peptide is released from the C-terminus of prosystemin, the 200 amino acid precursor, following post-translational modifications involving unknown events and enzymes. In tobacco, two systemin molecules have been recently isolated, neither sharing any sequence homologies with the tomato prosystemin gene/protein, but performing similar functions. We modified the tomato prosystemin gene by replacing the systemin-encoding region with a synthetic sequence encoding TMOF (trypsin-modulating oostatic factor), a 10 amino acid insect peptide hormone toxic to Heliothis virescens larvae, and expressed the chimeric gene in tobacco. The results reported here show that transformed leaves contain the TMOF peptide and exert toxic activity against insect larvae reared on them. In addition, subcellular localization studies showed the cytoplasmic location of the released TMOF, suggesting that in tobacco the enzymes responsible for the post-translational modifications of the tomato precursor protein are present and act in the cytoplasm to recognise the modified prohormone. The molecular engineering of the precursor, beside supplying new clues towards the understanding of prosystemin processing, constitutes an useful tool for plant genetic manipulation, by enabling the delivery of short biological active peptides.
番茄系统素是一种在受到伤害时产生的信号肽，它能在局部和全身激活多个防御基因。该肽是在经过涉及未知事件和酶的翻译后修饰后，从200个氨基酸的前体——前系统素的C末端释放出来的。最近在烟草中分离出了两种系统素分子，它们与番茄前系统素基因/蛋白质均无任何序列同源性，但功能相似。我们通过用编码TMOF（胰蛋白酶调节抑卵因子）的合成序列替换编码系统素的区域来修饰番茄前系统素基因，TMOF是一种对烟草天蛾幼虫有毒的10个氨基酸的昆虫肽激素，并在烟草中表达了嵌合基因。此处报道的结果表明，转化后的叶片含有TMOF肽，并对取食这些叶片的昆虫幼虫具有毒性活性。此外，亚细胞定位研究显示释放的TMOF定位于细胞质，这表明在烟草中存在负责番茄前体蛋白翻译后修饰的酶，且这些酶在细胞质中发挥作用以识别修饰后的激素原。前体的分子工程，除了为理解前系统素的加工提供新线索外，通过实现短生物活性肽的传递，还构成了植物基因操作的一种有用工具。
