AtMYB44通过调节内源H2S合成参与植物响应盐胁迫的分子机制

Hydrogen sulfide (H2S) is an important gaseous signal molecule. There are lots of reports presenting that the exogenous H2S participates in many plant physiological processes, especially in environmental stress responses. However, the study about the endogenous H2S in stress responses is limited. We previously found that in Arabidopsis, the endogenous H2S, synthesised from L-/D-cysteine desulphydrase (LCD and DCD), displayed a higher level under the salt treatment. The H2S biosynthesis mutants (lcd, dcd and lcd/dcd) were more sensitive to salt stress. In addition, we found that the transcription factor myb44 mutant showed salt sensitive phenotype, decreased gene expression of AtLCD and AtDCD, less H2S content, and inhibited H2S synthetase activity in the mutant. Further study indicated that the MYB44 transcription factor can bind to the promoter of AtLCD and AtDCD. These data suggested that AtMYB44 might regulate plant salt stress response through H2S signaling. In the study, we plan to identify the MYB44-regulated key cis elements of AtLCD and AtDCD, and analyze the binding regulation by the salt stress in vitro and in vivo. This study would not only greatly enhance our understanding of the molecular mechanism of how MYB44 regulating AtLCD and AtDCD genes expression to affect endogenous H2S synthesis, but also lay theoretical basis for the endogenous H2S participating in salt stress response.

H2S是近几年发现的重要气体信号分子，外源H2S提高植物抗逆性方面已有很多报道，但是对植物在响应逆境过程中内源H2S合成调控机制的了解非常有限。我们前期研究显示，盐胁迫下来自L/D-半胱氨酸脱巯基酶（LCD和DCD）途径的H2S水平明显升高，H2S合成突变体（lcd、dcd、lcd/dcd）均对盐的敏感性增强；转录因子myb44突变体与前者表型相似，且AtLCD和AtDCD基因表达下调、内源H2S含量下降、H2S合成酶活性降低；利用酵母单杂交证明AtMYB44与AtLCD和AtDCD启动子互作。这些结果强烈暗示了AtMYB44通过H2S信号对植物耐盐能力的调节作用。本项目拟在以上基础上，继续深入鉴定AtMYB44调控AtLCD和AtDCD的关键顺式作用元件，从体内和体外试验证实两者的结合受到了盐胁迫的调控，并进一步探究其响应盐胁迫的机制，为解析内源H2S参与盐胁迫的作用机制提供依据。

土壤盐碱化是制约农作物生长的重要环境胁迫之一，是全球广泛关注的农业问题。因此探究植物响应盐胁迫的分子调控机理，进而通过基因工程手段提高农作物耐盐能力，具有重要的研究意义和应用价值。有报道外源H2S能够提高植物的耐盐性，但是对植物在响应盐胁迫过程中内源H2S合成调控机制还有待深入探究。本项目前期发现，盐胁迫下来自L/D-半胱氨酸脱巯基酶LCD和DCD途径的H2S水平明显升高，H2S合成突变体 (lcd、dcd、lcd/dcd) 均对盐胁迫敏感性增强；转录因子myb44突变体与H2S合成突变体表型相似，且LCD和DCD基因表达下调、内源H2S含量下降、H2S合成酶活性降低；利用酵母单杂交证明MYB44与DCD启动子互作。暗示MYB44通过H2S信号调控植物盐胁迫耐受能力。本项目在此研究基础上，研究了内源H2S在拟南芥响应盐胁迫过程中的作用，克隆并鉴定了MYB44、MYB77、LCD和DCD等基因及其启动子。利用酵母单杂交、EMSA、双荧光素酶系统和烟草瞬时转录激活实验证明转录因子MYB44和MYB77可以与DCD基因启动子结合，表明盐胁迫下，MYB44和MYB77作为DCD上游的转录激活因子，激活DCD基因的表达；进而促进内源H2S合成，降低ROS含量，减少细胞的氧化损伤，从而提高拟南芥抵御盐胁迫的能力。本项目研究成果系统深入的解析了“MYB44/MYB77→DCD→H2S”模块调控植物抗盐性的分子机制，同时，初步探究了H2S通过硫巯基化修饰ABA合成关键酶ABA2调控拟南芥盐胁迫响应的作用机理，揭示了植物应答盐胁迫下的生理、代谢及其信号转导通路，有助于发展调控植物抗盐性的技术手段和相关产品的开发，为作物抗逆品种选育和精准栽培提供分子靶标，具有较好的理论创新和应用潜力。