Meindl T, Boller T, Felix G
Friedrich Miescher-Institute, P.O. Box 2543, CH-4002 Basel, Switzerland.
Plant Cell. 1998 Sep;10(9):1561-70. doi: 10.1105/tpc.10.9.1561.
Suspension-cultured cells of Lycopersicon peruvianum respond with rapid medium alkalinization and a strong increase of a MAP kinase-like activity when treated with subnanomolar concentrations of the plant wound hormone systemin. Systemin fragments comprising the N-terminal 14 amino acids (syst1-14) or the C-terminal four amino acids (syst15-18), added singly or in combination, were inactive as inducers of these responses. Syst1-14 but not syst15-18 antagonized activity of intact systemin in a competitive manner. Likewise, intact systemin showed stimulatory, syst1-14 antagonistic activity, and syst15-18 showed no activity in leaf pieces of tomato (L. esculentum) plants assayed for the induction of ethylene biosynthesis. To study the molecular basis of perception, we extended the C-terminal end of systemin by a tyrosine residue and radioiodinated it to yield systemin-125I-iodotyrosine. In membrane preparations of L. peruvianum, this radioligand exhibited rapid, saturable, and reversible binding to a single class of binding sites. Binding showed a dissociation constant of approximately 1 nM, and binding of radioligand was efficiently competed by unlabeled systemin but not by syst15-18 or structurally unrelated peptides. Binding was also competed by the systemin antagonists syst1-14 and syst-Ala-17 (IC50 of 500 and 1000 nM, respectively). Thus, this binding site exhibits the characteristics expected for a functional systemin receptor. Based on these results, we propose a two-step mechanism for systemin action, with binding of the N-terminal part to the receptor as the first step and activation of responses with the C-terminal part as the second step.
秘鲁番茄悬浮培养细胞在受到亚纳摩尔浓度的植物创伤激素系统素处理时，会迅速使培养基碱化，并显著增强丝裂原活化蛋白激酶样活性。单独或联合添加包含N端14个氨基酸（syst1 - 14）或C端4个氨基酸（syst15 - 18）的系统素片段，作为这些反应的诱导剂均无活性。Syst1 - 14而非syst15 - 18以竞争性方式拮抗完整系统素的活性。同样，在检测乙烯生物合成诱导的番茄（L. esculentum）叶片中，完整系统素表现出刺激活性，syst1 - 14表现出拮抗活性，而syst15 - 18无活性。为了研究感知的分子基础，我们在系统素的C端末端延伸了一个酪氨酸残基，并对其进行放射性碘化，得到系统素 - 125I - 碘酪氨酸。在秘鲁番茄的膜制剂中，这种放射性配体与一类单一的结合位点表现出快速、可饱和且可逆的结合。结合显示解离常数约为1 nM，放射性配体的结合可被未标记的系统素有效竞争，但不能被syst15 - 18或结构不相关的肽竞争。系统素拮抗剂syst1 - 14和syst - Ala - 17（IC50分别为500和1000 nM）也能竞争结合。因此，该结合位点表现出功能性系统素受体预期的特征。基于这些结果，我们提出系统素作用的两步机制，第一步是N端部分与受体结合，第二步是C端部分激活反应。
