Pathogenicity Related Function Of Gene FoHFI1 In Fusarium Oxysporum F. Sp. Cubense

Fusarium wilt of banana caused by the soil-borne fungus Fusarium oxysporum f. sp. cubense(FOC) is one of the most destructive diseases of banana. Among them race 4can infect almost all current varieties of banana and induce the most destructive threats on banana product worldwide. However, the molecular mechanisms underlying the virulence and pathogenicity of this fungal pathogen are still poorly understood. Till now, there is no effective measure to control the disease. Studies on pathogenic genes of FOC4 will help to understand its pathogenesis, and thus to provide a theoretical basis for the development of control strategies of the disease.Based on our previous laboratory’s screening, we found a mutant with significantly reduced virulence, in which gene HFI1(Histone H2 A Functional Interator 1)is affected by the T-DNA insertion. Here the gene is named as Fo HFI1 in FOC4. In this study, to further clarify the the function of Fo HFI1 during the pathogenic process, we knocked out the gene in FOC4 through split marker homologous recombination. The phenotype and virulence of Fo HFI1 knock-out mutants were also studied. The main results are as follows:1. Knock out of Fo HFI1. Using PEG-mediated protoplast transtormation, Fo HFI1knock-out mutants were obtained.2 Fo HFI1 had effect on multiple phenotypes of FOC4. Compared with wild type strain XJZ2, the Fo HFI1 knock-out mutants had less inclusion in hypha, growed slowly, were impaired in aerial hyphae, reduced in hyphae diaphragm range, could not produced any conidia. q RT-PCR analysis showed that the expression levels of the gene Htf1, the gene coding nitrite reductase, the gene coding MAPK protein kinase, the genes Stua, the genes Wet A and the gene coding chitin synthesis down-regulated in the Fo HFI1 knock-out mutants.3. Fo HFI1 had effect on extracellular cell wall-degrading enzyme(CWDE) activity.Enzyme activity assay showed that the activity of polygalacturonase and cellulase of theFo HFI1 knock-out mutants reduced compared to the wild type. The expression levels of the CWDEs coding genes(genes coding glycoside hydrolase, endoglucanase, exoglucanase 1precursor, succinate-semialdehyde dehydrogenase, xylanase etc.) were down-regulated in the Fo HFI1 knock-out mutants.4. Fo HFI1 was required for the full virulence of FOC4. q RT-PCR analyses showed that the expression levels of Fo HFI1 significantly up-regulated during the pathogenic process, which indicating that Fo HFI1 played an critical role in the virulence of FOC4. The Fo HFI1 knock-out mutants could penetrate cellophane, however, they could not make further infection on the tomato fruits. Pathogenicity test on host banana plants revealed that Fo HFI1 knock-out mutants couldn’t induce the disease compared with the wild type strain.5. Fo HF11 had effect on infection of FOC4. Histopathological assay indicated that the Fo HFI1 knock-out mutants can’t infect the roots of banana plants. q PCR analyses revealed that the content of Fo HFI1 knock-out mutants in the banana root tissue was reduced as the extension of inoculation compared with the wild type.In summary, the gene Fo HFI1 is essential for physiology and pathogenecity of FOC4.Deletion of Fo HFI1 results in significant reduction of CWDEs activity. Mutants can’t infect the host roots and further cause defective invading and reduced virulence in FOC4.