植物模式识别受体激酶FLS2配体识别及其激活机制研究

发布时间：2018-05-05 05:07

本文选题：FLS2-flg22-BAK1 + 共受体　；参考：《清华大学》2015年博士论文

【摘要】：植物因其含有丰富的营养物质和水分而成为众多病原微生物觊觎的目标。然而,植物与动物不同,植物营固着生活,无法从环境中逃离又不具有特化的免疫细胞、神经系统以及体液循环系统。为了适应周围环境,植物逐渐进化出以细胞为单位的应答和交流系统。而植物免疫的第一道防线就是通过细胞质膜定位的模式识别受体(PRR)识别病原菌特异性的分子模式(PAMPs/DAMPs)。其中,LRR-RKs是PRR的主要成员。FLS2是典型的LRR-RK,也是模式生物拟南芥中发现的第一个模式识别受体,通过识别细菌鞭毛蛋白N端保守的22个氨基酸(flg22)而发挥重要的免疫作用。Flg22的结合会诱导FLS2和LRR-RK BAK1的异聚化,进而起始免疫信号。目前,已有很多有关FLS2识别flg22的研究报导,但缺乏相关的结构信息。更重要的是,BAK1蛋白在flg22诱导的FLS2信号激活通路中的功能尚不清楚。本研究利用昆虫细胞分泌表达FLS2和BAK1的胞外区,通过pull down实验和凝胶过滤层析体外重组了flg22诱导的FLS2-BAK1异源复合物,并解析FLS2-flg22-BAK1胞外区的晶体结构,阐明flg22诱导FLS2-BAK1异源二聚化的分子机制。FLS2通过其超螺旋结构内表面细长的凹槽来识别flg22,BAK1与复合物中FLS2的C端有广泛直接的接触面,并且Flg22的结合以及与BAK1的相互作用并没有引起FLS2的构象改变或寡聚化。除此之外,BAK1的N端帽子结构特异性的识别复合物中flg22的C端,生化和细胞实验证明BAK1对flg22 C末端的识别对于FLS2-BAK1异源二聚体的形成至关重要,揭示了BAK1的功能是作为FLS2共受体而非信号增强子。根据我们的工作,表明FLS2-flg22-BAK1异源复合物的形成是受体激活以及跨膜信号的分子开关。由于BAK1可以与多个PRR或LRR-RK形成配体依赖的异源二聚体,本研究为理解和研究他们的激活机制提供了理论和方法依据。同时,与生长信号复合物BRI1-BL-BAK1的研究工作比较,我们提出植物细胞外pH的改变参与调解植物发育和免疫信号之间的平衡。
[Abstract]:Plants are coveted by many pathogenic microorganisms because of their rich nutrients and moisture. However, plants are different from animals in that they are fixed in life and cannot escape from the environment without specialized immune cells, nervous systems, and humoral circulatory systems. In order to adapt to the surrounding environment, plants have evolved cellular response and communication systems. The first line of defense of plant immunity is the molecular pattern of PAMPs / DAMPs which recognizes pathogen specificity by the pattern recognition receptor (PRR) located on the cytoplasmic membrane. LRR-RKs is the main member of PRR. FLS2 is a typical LRR-RKand the first pattern recognition receptor found in Arabidopsis thaliana. By recognizing 22 conserved amino acids of bacterial flagellin, flg22), the binding of Flg22 can induce the heteropolymerization of FLS2 and LRR-RK BAK1 and initiate the immune signal. At present, there are a lot of research reports about FLS2 recognition flg22, but lack of relevant structure information. More importantly, the function of flg22-induced FLS2 signal activation pathway is unclear. In this study, the extracellular domains of FLS2 and BAK1 were secreted by insect cells. The heterologous complexes of FLS2-BAK1 induced by flg22 were recombined by pull down experiment and gel filtration chromatography in vitro, and the crystal structure of the extracellular domain of FLS2-flg22-BAK1 was analyzed. The molecular mechanism of FLS2-BAK1 heterodimerization induced by flg22. FLS2 recognizes the wide and direct contact surface between flg22 and the C terminal of FLS2 in the complex by means of thin and long grooves on the inner surface of the superhelix structure. The binding of Flg22 and its interaction with BAK1 did not result in conformation change or oligomerization of FLS2. In addition, the N-terminal cap of BAK1 recognized the C-terminal of flg22 specifically in the complex. Biochemical and cellular experiments showed that the recognition of flg22 C-terminal by BAK1 was very important for the formation of FLS2-BAK1 heterodimer. It is revealed that BAK1 functions as a FLS2 coreceptor rather than as a signal enhancer. According to our work, it is suggested that the formation of FLS2-flg22-BAK1 heterologous complexes is the molecular switch of receptor activation and transmembrane signal. Since BAK1 can form ligand-dependent heterodimers with multiple PRR or LRR-RK, this study provides a theoretical and methodological basis for understanding and studying their activation mechanisms. At the same time, compared with the study of growth signal complex (BRI1-BL-BAK1), we suggest that the change of extracellular pH in plants is involved in regulating the balance between plant development and immune signals.
【学位授予单位】：清华大学
【学位级别】：博士
【学位授予年份】：2015
【分类号】：Q943.2

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