拟南芥GCR2参与感应N-丁酰基高丝氨酸内酯过程的初步研究

Preliminarily Research on Involvement of Arabidopsis GCR2 Responsing to N-Butyryl-DL-homoserine Lactone

N-丁酰基高丝氨酸内酯(C4-HSL)是革兰氏阴性菌主要的群体感应信号,其能促进植物生长发育,激活细胞膜表面的Ca2+通道,从而参与调控植物的生理代谢。然而,植物感应C4-HSL的分子机制并不清楚。拟南芥GCR2作为ABA的受体,对植物的生理代谢十分重要。旨在探寻GCR2是否参与拟南芥感应C4-HSL的过程。q RT-PCR结果表明,C4-HSL处理后1 h,GCR2基因表达量出现明显上调并在6 h后达到最大值,说明C4-HSL可调节GCR2。ELISA结果显示,GCR2蛋白表达量也在6 h达到最大值。对体外表达的GCR2进行纯化和浓缩,使其达到0.6 mg/m L后进行微量热泳动(MST)检测。MST测得C4-HSL与GCR2的解离常数(Kd)为166 nmol/L,显示出较强的结合能力。用BSA作为阴性对照,表明C4-HSL与GCR2的结合具有一定的特异性。这些结果表明GCR2可能参与了拟南芥感应C4-HSL的过程。

N-Butyryl-DL-homoserine lactone （ C4-HSL ） is a main quorum sensing signal in gram-negative bacteria. It could significantly promote root elongation and activate Ca2＋ channel at eytomembrane. C4-HSL can regulate metabolization of plant. However, little is known about the molecular mechanism of plants responding to C4-HSL. Arabidopsis thaliana GCR2 is receptor of abscisic acid （ ABA ） . It is important for metabolization of plant. This research aimed to explore whether the GCR2 is inyolved in the process of Arabidopsis thaliana reacting to C4-HSL. qRT-PCR showed that C4-HSL could regulate expression of GCR2. Expression of GCR2 was significantly upregulated after 1 h treated by C4- HSLand maximaized at 6 h. ELISA also showed that exPression of GCR2 maximaized at 6 h. GCR2 was purified and condensed to 0.6 mg/mL for Microscale Thermophoresis （ MST ） measurement. MS T indicated that dissociation constant （ Kd ） of GCR2 and C4-HSL was 166 nmol/L, which meant that they had strong binding affinity. Taking BSA as negative control, this certified that binding of GCR2 and C4-HSL had special character. These results showed that Arabidopsis GCR2 maybe involved in the pathway of plant responding to C4-HSL.