A Preliminary Study On The Functions Of Four Genes Regulated By CsTup1 In Bipolaris Sorokiniana

In the previous study,our laboratory conducted a preliminary study on the function of Tup1,a transcriptional suppressor in Bipolaris sorokini ana.The deletion of Tup1 gene led to the defect of growth and development,the absence of conidia and the decrease of pathogenicity.We analyzed the tran scriptome of the ΔCsTupl and found that many genes in the ΔCsTup1 mutant were differentially expressed compared with the wild type.In this study,four down-regulated genes were selected,including COCSADRAFT 157558,COCSA DRAFT₉0297,COCSADRAFT 194504 and COCSADRAFT₁16197.These fou r genes are conserved in the genus Bipolaris,and there are no reports in other organisms.In this study,the functions of these four genes were studied by ho mologous recombination technique,in order to find new pathogenic genes regul ated by transcriptional inhibitor Tup1.The main results are as follows:（1）COCSADRAFT₁57558 protein domain was predicted by SMART,and it was found that it had no typical functional domain.Through the study of biological phenotype,it was found that there was no significant difference in conidia morphology,appressorium production and pressure resistance between the ΔCs157558 mutant and the wild type,but the conidia production of the ΔCs157558 mutant decreased.Three sporulation-related genes StuA,MedA and WetA were significantly down-regulated in the mutant.The pathogenicity of Cs157558 to barley leaves decreased significantly.（2）COCSADRAFT 90297 contains a helix-loop-helix domain（HLH）,It was found that the colony morphology,conidia morphology,appressorium production,pressure resistance and pathogenicity to barley leaves of the ΔCs90297 mutant were not significantly different from those of the wild type,but the sporulation was significantly increased.Through the verification of the qRT-PCR of four sporulation-related genes StuA,MedA,WetA and VosA,it was further demonstrated that COCS ADRAFT 90297 was involved in the regulation of sporulation.（3）COCSADRAFT₁94504 contains a cytochrome b5-like heme/steroid binding domain,It was found that compared with the wild type,the ΔCs194504 mutant could not produce conidia,the mycelium grew very slowly,the mycelium became wider and deformed,and the expression of four chitin synthase related genes in the deletion mutant was significantly increased,and the resistance to oxidative stress and salt stress was enhanced.The resistance to osmotic stress weakened,while the pathogenicity of knockout mutants to barley leaves decreased significantly.In summary,COCSADRAFT₁94504 may be involved in sporulation,mycelium growth,pressure stress and pathogenicity regulation.（4）COCSADRAFT₁16197 contains an epidermal growth factor-like domain（EGF）,It was found that there was no significant difference in colony morphology,conidia morphology and yield,appressorium production and pathogenicity between the ΔCs116197 deletion mutant and the wild type,but the tolerance to oxidative stress was stronger than that of the wild type,indicating that COCSADRAFT 116197 may be involved in regulating the oxidative stress process of Bipolaris sorokiniana.To sum up,through the preliminary study on the function of these four genes,we found that COCSADRAFT₁57558,COCSADRAFT₉0297 and COCSADRAFT 194504 could affect the conidia production of Bipolaris sorokiniana in wheat root,indicating that these three genes may be involved in the sporulation process of fungi.In addition,COCSADRAFT₁57558 and COCSADRAFT₁94504 also affected the pathogenicity to barley leaves.The results showed that COCSADRAFT₁57558 and COCSADRAFT₁94504 were also involved in regulating the pathogenicity of Bipolaris sorokiniana to barley and wheat leaves.Although COCSADRAFT 116197 is not involved in the regulation of sporulation and pathogenicity,it is likely to be involved in the process of oxidative stress.