转录因子CfHac1调控油茶果生刺盘孢响应内质网压力和致病力分子机制研究

果生刺盘孢是油茶炭疽病的优势流行致病菌，但其致病的分子机制尚不清楚。我们前期研究发现果生刺盘孢侵染油茶或受到二硫苏糖醇胁迫时，转录因子CfHAC1基因通过可变剪切产生了2种特异的mRNA转录本。因此，我们提出CfHAC1基因通过可变剪切产生 2 种特异的转录本来响应寄主或二硫苏糖醇造成的内质网压力胁迫假说。本项目拟分别构建果生刺盘孢CfHAC1基因常规剪切和2种特异剪切共3种转录本的表达载体，采用ChIP-Seq技术鉴定它们在病菌中结合的DNA序列，采用定量PCR并结合课题组已有转录组数据筛选目标基因，采用EMSA和Y1H验证鉴定3种转录本的靶基因，并分析靶基因的生物学功能。比较分析2种特异转录本所调控的基因，解析它们在响应内质网压力胁迫和致病过程中的功能。最终验证我们的假说并阐明CfHac1调控果生刺盘孢响应内质网压力胁迫和致病的分子机制。研究结果对油茶炭疽病防控具有重要指导意义。

Anthracnose on Camellia oleifera is one of the most serious diseases of Camellia in China. The major causal pathogen of Camellia anthracnose is Colletotrichum fructicola, but the detailed molecular mechanisms of its pathogenicity is still largely unclear. In previous study, we characterized the bZIP transcription factor CfHac1 control the development and pathogenicity in C. fructicola. Our formal study also revealed that here are two special alternatively spliced transcripts of CfHAC1 gene to respond to endoplasmic reticulum(ER) stress in C. fructicola. This study will construct the expression vectors of these three alternatively spliced transcripts of CfHAC1 gene, and identify their DNA binding sequences in the pathogen using ChIP-seq(Chromatin immunoprecipitation high-throughput sequencing). In combination with transcriptome data and quantitative PCR analysis, the target genes of three alternatively spliced transcripts of CfHAC1 will be screened. Electrophoretic mobility shift assay and yeast one hybrid will be employed to identify the target genes. Study of the target genes biological function will be performed by gene knockout. To elucidate the molecular mechanism of two special alternatively spliced transcripts factors in regulating endoplasmic reticulum stress response and pathogenicity in C. fructicola, the expression of the two special alternatively spliced CfHAC1 transcripts regulated genes will be elucidated. This research not only provide the theories of revealing the molecular mechanism of CfHAC1 in response to endoplasmic reticulum stress and pathogenicity, but also has important guiding significance to discover novel fungicide targets and improve management methods to control disease.