| Northern Leaf Blight of Corn, caused by Setosphaeria turcica, is an very important fungal disease, which resulted in serious economic losses on the production of corn. It was current studies found that, cAMP, MAPK and Ca2+ signal transduction pathway mediated by G protein played an important role in plant pathogenic fungi growth, development and disease in many aspects. Type 2A phosphoprotein phosphatase(PP2A) which located in the downstream signal transduction pathways was a major Ser/Thr phosphatase. PP2 A was involved in the regulation of signal transduction pathways with various protein kinase by the phosphorylation and dephosphorylation.The deletion mutant of StPP2A-c gene had been acquired through gene knockout technology in our laboratory. It was found that, St PP2A-c proformed a negative regulation of conidia and melanin synthesis in the preliminary study. In order to further clarify the gene function and regulation mechanism, we constructed the response vector of StPP2A-c gene, obtained the revertant of StPP2A-c gene through the transformation of protoplasts, and analyzed the transcriptome of the wild type strain and the StPP2A-c gene deletion mutant. In addition, to analyze the relationship between PP2 A and the signal teansduction pathways, the expression levels of genes of MAPK kinase in MAPK signal pathway and calmodulin(CAM), calcineurin(CaN) and calmodulin dependent protein kinases(CaMKs) in Ca2+ signal pathway by the Real-time PCR in â–³StPP2A-c deletion mutant. The results were as follows:1.In S. turcica, St PP2A-c gene proformed the negative regulation in formation of conidia and the synthesis of melanin. And lack of this gene also resulted in obviously lagging behind of the conidial germination and appressorium formation, and was more sensitive to oxygen stress.2. By the transcriptome sequencing of the wild type strain and the â–³StPP2A-c mutant, it was found that the deletion of StPP2A-c gene leaded to the obvious difference in the expression of 791 genes, in which 323 genes were significantly down regulated, 468 genes were up-regulated. And most of these genes encoded the unknown function proteins and the rest belongs to the glycosyl hydrolase family and transferase family, which mainly involved in the regulation of tryptophan metabolism pathway.3. Using bioinformatics methods,it was finally confirmed that there were four CaMK genes in the genome of S. turcica, named CAK1, CAK2, CAK3 and CAK4. Relative expression quantities of CaMK genes in the invasive structure formation process were detected by using the Real-time PCR technology. Transcriptional pattern analysis showed that CAK2 and CAK3 were actively transcribed, and the transcriptional levels in every stage were higher than that at 0 h post induction(hpi). Especially, the expression levels of both CAK2 and CAK3 were up-regulated at 6 and 24 hpi, significantly(P<0.05). On the contrary, the transcription levels of CAK1 and CAK4 in all stages were significantly lower than the mock(P<0.05).4. In wild type, â–³StPP2A-c mutant and revertant, relative expression quantities of the genes of MAPK kinases(STK1, STK2, STK3), calmodulin(CAM), calcineurin(CaN), calmodulin dependent protein kinases(CaMKs) were detected by Real-time PCR technology. It is found that, the deletion of St PP2A-c gene had no effect to the expression of CAK3 and CAK4 gene of CaMKs, while the expression of other genes were up-regulated, in which STK1, CAM, CNB, CAK1 and CAK2 showed significant increase(p<0.05). |
