数字基因表达谱及混合模型研究稻瘟病菌细胞自噬与相关通路基因互作

Magnaporthe oryzae is pathogenic to a number of economically important crops such as rice, barley, wheat, and millet. It utilize a specially developed complex infection structure, the appressorium, to gain entry to and colonize plant tissue. The appressorium generates immense intracellular turgor pressure allowing it to penetrate the leaf cuticle. This enormous turgor was a consequence of the accumulation of huge quantities of glycerol in the cell. A number of studies have revealed autophagy is responsible for this process. The core components of the autophagy machinery have been cloned and analyzed. Similar with the progress of autophagy research in other model organisms, such as Saccharomyces cerevisiae and Homo sapiens, the over-all regulatory network of autophagy and the underlay mechanism remained unknown. Until recently, there was a publication described a snap shot of the basal-level autophagy network in H. sapiens. Development of the high-throughput sequencing technology and achievements in statistical genetics made the direct evaluate of gene-gene interaction and transcriptional co-regulation a reality. This program focus on the analysis of autophagy related gene-gene interaction of ΔMoATG8 and the relative widetype under autophagy-induced and normal conditions. By utilizing the powerful mixed-linear model approach, a number of epistasis can be predicted between the different pairs of differentially expressed genes (DEGs). And the related pathway information can be extracted by comparative study with S. cerevisiae and H. sapiens. It is expected to make the autophagy regulatory network during appressoria development in this fungus and the transcription-level co-regulatory mechanism clear, as well as new components of the autophagy process. All these efforts will make the antifungal drug development become true.

稻瘟病菌的侵染能力与附着胞正常发挥功能呈正相关。附着胞正常发挥作用依赖于胞内具有的巨大膨压，已知这种膨压的产生与细胞自噬过程密不可分。目前稻瘟病菌中的核心自噬基因已基本得到了分离与初步鉴定。但和自噬过程总体研究情况类似，这样的鉴定只限于一些核心蛋白复合物各组分的探讨，而缺乏对于自噬过程整体上的把握以及与该过程相联系的其它信号通路cross-talking情况的了解。高通量测序技术和统计遗传学方法的发展使得对稻瘟病菌自噬过程基因互作状况以及转录水平上的协同调控研究成为现实。本项目拟利用RNA-seq和tag profiling技术获得野生型和△MoATG8突变体在自噬诱导/正常条件下的表达谱，并利用混合模型和生物信息学手段获得差异表达基因之间的互作信息及通路分布情况，初步阐明稻瘟病菌附着胞发育的自噬基因互作网络和转录水平的协同调控机制，寻找自噬过程的新组分。为今后药物靶标筛选提供理论依据。

稻瘟病菌的侵染能力与附着胞正常发挥功能呈正相关。附着胞正常发挥作用依赖于胞内具有的巨大膨压，已知这种膨压的产生与细胞自噬过程密不可分。目前稻瘟菌中的核心自噬基因已基本得到了分离与初步鉴定。本项目利用RNA-seq和tag profiling技术获得野生型和△MoATG1突变体在自噬诱导/正常条件下的表达谱，并利用混合模型和生物信息学手段获得差异表达基因（DEGs）之间的互作信息，初步阐明稻瘟病菌附着胞发育的自噬基因互作网络和转录水平的协同调控机制，寻找自噬过程新组分。. 目前，完成了酵母和人中自噬相关的PPI和通路信息的整理，使用graph相关的方法构建了可靠的ATGs的PPI互作网络，含有~500 unique的PPIs；完成了附着胞发育0h、8h、12h、18h、24h的表达谱测定和分析，发现DEGs在附着胞的不同发育阶段分别具有11和13种显著的基因表达模式，特异性较高。功能分析表明，DEGs在cellular ion homeostasis, ion transport, lipid metabolic process, lipid transport, monocarboxylic acid metabolic process, protein glycosylation, sporulation, carbohydrate metabolic process, carbohydrate transport中富集；进一步的分析表明下调基因主要集中在lipid metabolic process, amino acid transport, protein glycosylation三个通路中，上调基因则分散分布在36个功能/通路中，没有发生明显富集；在发育模式为显著负相关的7个DEGs挑选了3个可能与MoATG1存在直接互作的基因进行功能验证。研究产生的测序数据已递交NCBI，部分重要结果也建立了可视化查询网站，方便合作者查询。. 根据最新的NCBI公共数据库查询，本研究是稻瘟菌中最大规模的系统化发育信息研究，对附着胞发育过程的研究和新方向的开辟必然产生较大的影响；执行中碰到的基因组和转录组的复杂性改变了之前对稻瘟菌的一些看法；同时，为后续研究的开展和类似项目的大规模筛选提供了技术支持和实施经验。