TaFKBP65调控小麦耐热性的分子机理研究

As global temperature continues to rise, high temperature will seriously affect the yield and quality of wheat. Thermotolerance of wheat is a quantitative trait controlled by multiple genes, and due to the lack of an intuitive and reliable evaluation index, it is difficult to breed new thermotolerant wheat varieties by conventional methods. Therefore, identification of thermotolerant genes and studying their molecular mechanism are of great significance to the molecular breeding of new thermotolerant wheat varieties. Recent studies demonstrated that AtFKBP62 (ROF1) and AtFKBP65 (ROF2) are involved in long term acquired thermotolerance through its interaction with Hsp90 and modulation of the heat shock transcription factor HsfA2, thus further affect small heat shock proteins accumulation during the recovery period and ultimately regulating acquired thermotolerance development. However, the functional studies and molecular mechanism of wheat FKBP under heat stress is remain unknown. We analyzed the proteome of wheat variety “Chinese spring” by iTRAQ method, and found three TaFKBP65s (TaFKBP65_2AL, TaFKBP65_2BL, TaFKBP65_2DL) were significantly up-regulated under heat stress. Homologous analysis of amino acid sequence showed that the three TaFKBP65s are highly homologous to AtFKBP62 (ROF1) and AtFKBP65 (ROF2) from Arabidopsis. In this study, we will clone the three TaFKBP65 genes and verify their gene function resistant to heat in Arabidopsis and wheat by transgenic technology, meanwhile reveal the molecular mechanism of TaFKBP65 resistant to heat by RNA-seq analysis. It will provide a good theoretical foundation for breeding new thermotolerant wheat varieties.

高温严重影响小麦的产量和品质。由于小麦耐热性是受多基因控制的数量性状且缺少直观、可靠的评价指标，通过常规方法很难选育耐热新品种。因此，克隆鉴定耐热基因并研究其分子机理，对耐热性小麦分子育种具有重要意义。近期研究表明，拟南芥FKBP类蛋白AtFKBP62和AtFKBP65能与热激蛋白HSP90相互作用共同调节热激转录因子HsfA2的转录活性，调控植物在高温胁迫下的生长发育。申请人使用iTRAQ技术分析小麦在高温胁迫下蛋白组的变化，共发现3个TaFKBP65显著上调表达。氨基酸序列同源性分析表明3个TaFKBP65与拟南芥AtFKBP62、AtFKBP65高度同源。本项目将克隆以上3个TaFKBP65基因，通过转基因技术鉴定它们的耐热功能，采用RNA-seq技术揭示TaFKBP65调控小麦耐热性的分子机理，为培育耐热性小麦新品种奠定理论基础。

高温严重影响小麦的产量和品质。由于小麦耐热性是受多基因控制的数量性状且缺少直观、可靠的评价指标，通过常规方法很难选育耐热新品种。因此，克隆鉴定耐热基因并研究其分子机理，对耐热性小麦分子育种具有重要意义。本项目通过克隆热响应基因TaFKBP65全长序列和启动子序列，小热激蛋白TaHSP23.9基因全长序列，TaRAD23基因全长序列，研究这些基因在调控小麦耐热性方面的功能。结果表明，这三个基因均受热显著上调表达，将这三个基因在拟南芥中过量表达，发现能提高转基因拟南芥的耐热性，而且过表达TaHSP23.9还能提高转基因拟南芥的耐盐性。此外，对TaFKBP65启动子序列分析发现，该启动子区包含多种顺式作用元件，例如热激转录因子特异结合的“SHE”元件，参与光信号转导的“G-box”元件，参与脱落酸信号转导的“ABRE”元件等。同时，GUS染色分析表明TaFKBP65启动子能显著被热诱导并促下游基因的表达，而且该启动子可能是一类组成型启动子，在检测的多个组织中均有表达。这些研究结果将有助于我们理解小麦耐热性的分子机制，同时克隆到的热诱导型启动子和耐热基因为今后通过转基因方法获得小麦耐热新品种提供理论基础。