绿僵菌是一类应用于生防的昆虫病原真菌，孢子耐热差限制了其大田应用效率和商品化程度。我们近期发现：在罗伯茨绿僵菌孢子热胁迫过程中存在数千上调基因，经pathway分析显著富集于内吞途径。在真核生物中，网格蛋白介导的内吞途径（CME）直接调控胞内外信号传递，反之，CME也可被actin等信号通路调控；因而在逆境胁迫应答等过程中起重要作用。但鲜有昆虫病原真菌的相关研究报道。本项目在前期基础上，采用基因敲除/回补和激光共聚焦显微观察等技术，分析绿僵菌Chc（CME的关键基因）和Ark1（模式真菌中，作用于actin信号通路调控CME）基因对孢子CME及耐热力的影响，从而明确CME对孢子耐热力的作用；结合深度测序和定量磷酸化蛋白质组等手段鉴定孢子耐热中CME调控的信号通路及actin信号通路调控CME的关键分子，阐明CME在孢子耐热中的作用机理；为真菌杀虫剂田间适应性和稳定性的遗传改良提供理论依据。

Metarhizium is an insect pathogenic fungi widely used as a biocontrol agent, low thermotolerance of conidia is the main impediment to the efficiency of field applications and the level of commercialization. In our preliminary studies, thousands of up-regulated genes were identified by differential expression analysis at the transcriptome level in heat-treated conidia, and the results of KEGG pathway enrichment analysis show the endocytosis pathway was most significantly enriched pathway for up-regulated genes. In Eukaryote, Clathrin-mediated endocytosis(CME) has many effects on signal transduction and, conversely, actin signalling can regulate CME; and therefore it play an important roles in the stress response. However, little is known about related research for entomopathogenic fungi...Based on the preliminary work, firstly, functions of gene Chc(key gene in CME) and Ark1(that control CME through regulating actin signalling in model fungi) will be analyzed and identified by gene deletion/complementation and Confocal microscopy, therefore effects on the conidial thermotolerance of CME will be confirmed. Secondly, experimental methods,such as high-throughput sequencing and quantitative phosphoproteomics, will be employed to identify signaling pathways and key proteins in the intertwining molecular networks of CME and actin signalling, and therefore the regulatory mechanism of CME in thermotolerance of Metarhizium conidia will be clarified.The findings should provide a theoretical basis for the genetic improvement of the production strain of Metarhizium.