In phytopathogenic fungi, the Hog1 MAPK pathway plays roles in osmoregulation, oxidation responses, fungicide sensitivity, among others. However, how fungi depend on the HOG MAPK pathway to regulate their functions is poorly understood. To further elucidate the molecular mechanisms of the HOG MAPK pathway, a yeast two-hybrid (Y2H) system was adopted to screen interacting proteins of two key components (CsPbs2 and CsHog1) of the HOG MAPK pathway in a cDNA library of Colletotrichum siamense. The results showed that 19 proteins may interact with CsPbs2, including cytochrome P450 4F5, ceramide very-long-chain fatty acid hydroxylase SCS7, the oxidative stress response two-component system protein SSK1, and other proteins. Twenty-one proteins may interact with CsHog1, including the cell wall protein PhiA, acriflavine sensitivity control protein acr-2, arachidonate 15-lipoxygenase, respiratory supercomplex factor 2-like protein, and others. Five proteins interact with both CsHog1 and CsPbs2: antigenic thaumatin-like protein, aspirochlorine biosynthesis protein N, secreted beta-glucosidase sun 1, alcohol dehydrogenase 1, and hypothetical protein GQ607_005106. Furthermore, the interaction of CsSCS7 and CsCytP450 4F5 with CsPbs2 was verified by His pull-down. qPCR revealed significantly upregulated CsPbs2, CsHog1, CsSCS7 and CsCytP450 4F5 expression in Colletotrichum treated with 50 μg/mL fludioxonil for 2 h. This research identified candidate CsHog1- and CsPbs2-interacting proteins via a Y2H system and confirmed the interactions of CsSCS7 and CsCytP450 4F5 with CsPbs2. The findings lay the foundation for further studies of HOG MAPK pathway function and regulatory mechanisms, especially regarding fungicide sensitivity.

在植物致病真菌中，Hog1 MAPK途径在渗透调节，氧化反应，杀菌剂敏感性等方面发挥作用。但是，人们对真菌如何依赖HOG MAPK途径调节其功能的了解甚少。为了进一步阐明HOG MAPK途径的分子机制，采用酵母双杂交（Y2H）系统在暹罗Colletotrichum siamense cDNA文库中筛选了HOG MAPK途径的两个关键成分（CsPbs2和CsHog1）的相互作用蛋白。结果表明，19种蛋白质可能与CsPbs2相互作用，包括细胞色素P450 4F5，神经酰胺超长链脂肪酸羟化酶SCS7，氧化应激反应两组分系统蛋白质SSK1和其他蛋白质。21种蛋白可能与CsHog1相互作用，包括细胞壁蛋白PhiA，a啶黄敏感性控制蛋白acr-2，花生四烯酸15-脂加氧酶，呼吸超复合因子2样蛋白等。五个蛋白与CsHog1和CsPbs2相互作用：抗原类索马甜蛋白，阿司匹林生物合成蛋白N，分泌的β-葡糖苷酶sun 1，醇脱氢酶1和假定蛋白GQ607_005106。此外，通过His下拉证实了CsSCS7和CsCytP450 4F5与CsPbs2的相互作用。qPCR显示CsPbs2明显上调，CsHog1，CsSCS7和CsCytP450 4F5表达炭疽病用50μg/ mL的咯菌腈处理2小时。这项研究通过Y2H系统鉴定了与CsHog1和CsPbs2相互作用的候选蛋白，并证实了CsSCS7和CsCytP450 4F5与CsPbs2的相互作用。这些发现为进一步研究HOG MAPK通路功能和调节机制奠定了基础，尤其是在杀菌剂敏感性方面。