Functional Characterization Of Fusarium Graminearum TEP1 Gene

Fusarium graminearum is the major causal agent of Fusarium head blast (FHB) disease on wheat, The infection process of F. graminearum in host plants includes conidia production and surface attachment,conidia germination,infectious hypha differentiation and spreading.All these stages are common in many other plant pathogenic fungi.Therefore,studying on the molecular basis of these developmental stages in F. graminearum will help to elucidate the molecular mechanism of fungal pathogenicity and to discover candidate target genes for screening and designing novel fungicides.In recent years, F.graminearum was carried out a great deal of studies in the world and made a series of significant progress. However, due to toxins in the infection process, making it so far on the pathogenesis of F.graminearum and resistance mechanisms of host is still a lack of comprehensive understanding and awareness.At present,the studies on F. graminearum have been focused on the pathogenesis and the discovery of drug targets. In this study,a novel gene FgTEP1 required for conidiation was identified and linked to the phosphatidylinositol pathway and the pathogenicity of F. graminearum.The results in details are as the following:Firstly,we constructed the knock-out plasmid pMDa+b+G418 for the FgTEP1 gene and successfully deleted the ORF of FgTPE1 gene by homologous recombination. Compared with wild-type strain PH-1, the knock-out strain Fgtep1-t are sensitive to both lithium ions and phosphatidylinositol-3-phosphate(PI3)kinase inhibitor wortmannin,the conidia production of Fgtep1-t is less than 56% and its virulence is less about 38.5%.To further validate the function of FgTPE1 gene,we constructed the complemental plasmid pUC-hph-FgTEP1 for gene complement to Fgtep1-t.The gene complement results showed that all the capacities of the resistance to wortmannin and to lithium ions,the conidia production and the pathogenicity in the complementary transforments were restored to the level of those in the wild-type strain PH-1 by 3.4kb DNA fragment,including functional promoter and the entire coding region of the FgTPE1.These results proved that the mutant phenotype was caused by the detection of FgTPE1.This study also exploits the possible complementary function of the FgTEP1 gene in Saccharomyces cerevisiae. We cloned the cDNA sequence of the FgTEP1 gene into a yeast expression vector YPB1-ADH1 with a strong promoter of ADH1, the alcohol dehydrogenase gene. As a result, we have found that the FgTEP1 cDNA sequence could complement the function of ScTEP1 in wortmannin tolerance, which indicates that the FgTEP1 and ScTEP1 genes are functional homologues and linked to the phosphatidylinositol pathway.This study provides a basis for further studies on the phosphatidylinositol pathway, conidia production and virulence of Fusarium graminearum. It also provides an important guiding significance for the development of a new type of broad-spectrum fungicide.