花生旱后恢复生长因子AhPGRF1的功能研究

The crop yield and quality of peanut could be significantly restrained by drought. However, the study on peanuts growth recovery after drought is rare and the regulating mechanism is not clear at present. Recently, a GLK-like transcription factor has been screened from the yeast cDNA library by AhHDA1. Peanut leaves become wilting together with a rapid decline of the gene expression under drought treatment during 72 to 96 h. The expression of GLK-like gene was gradually increasing after rehydration, and the expression of which promoted the transcription activity of AhPORA, an key gene of chlorophyll synthesis . The GLK-like gene was renamed as AhPGRF1 (Arachis hypogaea Post-Drought Growth Recover Factor 1). On the basis of this, this project intends to study the function of AhPGRF1 gene expression and protein expression in the process of growth recovery after drought. It will be understand the relationship between AhPGRF1 with histone acetylation .The transcription regulation mechanism of histone acetylation modification of AhPGRF1 and the effect of growth recovery after drought mediated by AhHDA1 will be studied. The results will provide basis to uncover the molecular mechanism of growth recovery after drought by AhPGRF1, and then an understanding of the mechanism growth recovery after drought will be promoted.

干旱严重制约花生的产量与品质。目前对花生在旱后恢复生长的研究甚少，调节机制不清楚。课题组前期从花生去乙酰化酶的酵母库中筛选克隆出一个GLK类转录因子基因。花生在干旱下该基因表达快速下降，复水恢复生长后表达逐渐上调，其影响叶绿素合成关键酶AhPORA基因启动子转录活性，命名为花生旱后生长恢复因子1（AhPGRF1）。在此基础上，本项目拟研究花生旱后恢复生长过程中AhPGRF1基因和蛋白表达的作用，明确AhPGRF1与组蛋白去乙酰化的相互关系, 探讨AhPGRF1与AhHDA1互作位点及其复合体在干旱及旱后恢复生长过程中的动态变化，以及对靶基因（AhPORA）的转录调节，阐明AhPGRF1与组蛋白乙酰化修饰的关系及其在花生旱后恢复生长中的功能。研究结果为揭示AhPGRF1调节花生旱后恢复生长的分子机制提供依据，促进对作物旱后恢复生长机制的深入了解。

在前期从花生 cDNA 文库中筛选到转录因子 AhGLK1的基础上，探讨AhGLK1 在花生旱后恢复中的作用以及调节花生旱后恢复生长的分子机制。研究结果表明，花生叶片在干旱时，其叶绿素含量与荧光参数、AhGLK1基因和蛋白表达、光合作用基因（AhLHCB3 和 AhRuBp)和叶绿素合成相关基因（AhPORA、AhCHLH、AhHEMA和 AhCAO)的表达降低，复水恢复后升高。拟南芥AhGLK1/glk1glk2 的旱后生存率显著高于glk1glk2株系, 其叶色恢复绿色，叶绿体发育正常，叶绿素含量与荧光参数提高，叶绿素合成相关基因和光合作用相关基因（LHCB2.1、LHCB3 和 LHCB6）的表达水平皆显著提高。AhGLK1-OX 花生毛状根呈绿色，有大量叶绿体。AhHDA1能结合到AhGLK1启动子区域，改变组蛋白去乙酰化水平从而抑制AhGLK1的表达。RNA-seq证实叶绿素合成和光合作用相关基因的表达在AhGLK1超表达毛状根中升高，而在AhGLK1干扰毛状根中表达降低。进一步通过ChIP-seq发现了AhGLK1与AhHDA1共同的靶基因AhCAB。转录活性检测结果显示AhGLK1促进AhCAB的表达，而AhHDA1通过与AhGLK1互作抑制AhCAB和AhPORA表达。AhGLK1启动子P2区域是AhGLK1和AhHDA1关键结合位点。ChIP-qPCR结果显示在干旱和复水条件下，AhGLK1与AhHDA1结合AhCAB和AhPORA启动子区域调控其表达。我们推测在花生响应干旱和复水恢复生长时，AhGLK1-AhHDA1发挥开关作用调节AhCAB和AhPORA的表达，AhGLK1激活而AhHDA1抑制它们的表达, 影响着叶绿素合成和光合作用。