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激活蛋白PeaT1是从极细链格孢菌(Alternaria tenuissima)中分离的,能诱导植物产生系统获得抗性的蛋白激发子。为了阐明PeaT1诱导植物提高抗性的分子机制,本文采用RT-PCR技术研究了PeaT1诱导烟草悬浮细胞后不同时间PR-1a基因的转录活性,在烟草悬浮细胞中加入20μg/mL的PeaT18h后,PR-1a基因转录活性达到最高;相同浓度的PeaT1处理烟草植株,烟草叶片中酸性PR蛋白表达量升高。激光共聚焦显微镜观察到FITC荧光素标记的PeaT1可与烟草悬浮细胞表面结合;免疫荧光法进一步证明PeaT1可与烟草细胞质膜结合;使用共价交联剂BS3证明125I-PeaT1可与烟草细胞质膜上的2个分子结合,而与加热或蛋白酶处理的质膜不能发生交联,说明与PeaT1结合的分子具有蛋白特性,分子量分别为20kDa和30kDa。上述实验结果证明在烟草细胞质膜上存在着激发子PeaT1受体样的结合位点。
The activator protein PeaT1 is isolated from Alternaria tenuissima and can induce plants to produce systemically-acquired protein excitons. In order to elucidate the molecular mechanism of PeaT1-induced plant resistance, the transcriptional activity of PR-1a gene at different times after PeaT1 induction of tobacco suspension cells was studied by RT-PCR. After adding 20μg / mL PeaT18h to tobacco suspension cells, PR -1a gene transcription activity reached the highest; the same concentration of PeaT1 treatment of tobacco plants, tobacco leaf PR acid protein expression increased. Confocal laser scanning microscopy showed that FITC fluorescein-labeled PeaT1 could bind to the surface of tobacco suspension cells. Immunofluorescence confirmed that PeaT1 could bind to the plasma membrane of tobacco cells. The covalent cross-linking agent BS3 proved that 125I-PeaT1 binds to the plasma membrane of tobacco Of the two molecules, whereas the heat or protease-treated plasma membrane did not cross-link, indicating that the molecules bound to PeaT1 have protein characteristics with molecular weights of 20 kDa and 30 kDa, respectively. The above experimental results show that there is an elicitor PeaT1 receptor-like binding site on the plasma membrane of tobacco.