一个小麦ERF新基因的功能与分子机理研究

Wheat soil-borne disease， such as sharp eyespot and common root rot, are the most severe diseases that hazard wheat production. It is very necessary to study wheat defense mechanism to soil-borne disease. In the preliminary work, using Genechip and virus-induced gene silencing analysis, a novel wheat gene,temporally named TaERF-R1, encoding ethylene response factor, was identified to be involved in the wheat resistant to pathogene Rhizoctonia cerealis. Phylogenetic analysis suggests TaERF-R1 belongs to B-2 subgroup of ERF subfamily, members of which had few functional study on resistant responses. In this proposal, we plan to perform the following researches: to isolate the full-length sequences of the TaERF-R1 from various resistant and susceptible wheat cultivars and the progenitors of common wheat (T.urartu; Ae. Speltoides and Ae.tauschii); to ascertain TaERF-R1 transcription profiles and transcriptional activation activaty of protein production; to study the biological function and underlying mechnism of TaERF-R1 in wheat resistantce to soil-borne disease and abiotic stress through transgenic wheat plants overexpressing and underexpressing TaERF-R1. This project will provide the theorical basis for understanding the molecular mechanism of wheat defense responses, the resistant gene resource, and the thchnical assitance for wheat breeding with resistance against soil-borne disease ,such as sharp eyespot and common root rot in wheat.

小麦土传病害，如纹枯病和根腐病等已成为危害我国小麦生产的第一大病害。小麦抗土传病害分子生物学研究显得尤为重要。申请人所在的张增艳课题组前期利用转录组学技术、实时定量PCR，结合VIGS实验初步分析，鉴定出1个新的小麦抗纹枯病相关乙烯反应因子基因，暂时命名为TaERF-R1，进化树分析表明，该基因属于ERF亚家族的B-2组，该组成员目前很少有抗病相关功能报道。 本项目将从不同抗纹枯病和感病小麦及其二倍体祖先种中克隆该基因的全长序列，分析其转录表达特性和转录激活功能。通过转基因小麦创制、分子检测、表型鉴定，研究TaERF-R1在小麦抗纹枯病等土传病害和抗逆性中的生物学功能，进而以转基因植株为材料，利用转录组学、实时定量PCR、生物信息学和酵母单杂交等技术，研究其调控抗性反应的分子机理。该项目为深入认识小麦土传病害抗性分子生物学基础和抗性机理提供新线索，并为小麦抗性改良提供新的基因资源。

在前期研究中，通过转录组分析，分离到一个表达量在纹枯病侵染时显著上调的新型ERF转录因子基因TaERF-R1。降低CI12633中TaERF-R1的表达量可以削弱小麦对纹枯病的抗性。在此基础上，我们拟通过转基因小麦过表达TaERF-R1等方法进一步研究其在小麦抗纹枯病中的功能，并研究其作用机制。在本研究中，共得到多个过表达的扬麦16转基因株系，转基因小麦株系多代（T1-T3代）纹枯病和根腐病抗性鉴定发现，TaERF-R1的过表达并没有提高扬麦16对纹枯病的抗性。在结题报告正文中，对其可能的原因进行了分析。TaERF-R1的N端起始序列为MCGGAI，属于ERF VII类成员，符合拟南芥中响应低氧胁迫的RAP2.2、RAP2.12等ERF转录因子的结构特点。受此启发，研究发现，TaERF-R1的表达响应小麦湿害（水涝，waterlogging）胁迫。基因沉默和转基因过表达研究表明，TaERF-R1在小麦调节耐湿胁迫中具有重要作用。通过比较转录组的方法表明，TaERF-R1主要通过调控小麦的糖代谢进而调节小麦对湿害胁迫的耐性。目前正在对其可能的直接调控靶标进行验证。以上结果表明TaERF-R1是小麦调控湿害耐性的关键基因。该研究为改良小麦湿害耐性提供了理论基础和基因目标，所获得的转基因株系对于改良我国长江中下游麦区和西南麦区的小麦耐湿性具有重要作用。