Studies On The Functions Of Protein Disulfide Isomerase,PDI Gene Family In Pathogenicity Of Fusarium Oxysporum F.Sp.Niveum

Fusarium wilt disease is a common fungal disease of watermelon,which seriously threatens to the development of watermelon industry.This disease is caused by Fusarium oxysporum f.sp.niveum(Fon).The mechanim of the pathogenicity in Fon toward watermelon is largely unknown.This present paper is to explore the PDI(protein disulfide isomerase)gene function in Fon pathogenicity and their possible mechanisms.A total of six genes Fon00140,Fon08535,Fon01620,Fon08898,Fon18919 and Fon05843 were identified as members of the PDI family in Fon and confirmed by cloning and sequencing.Transformants were obtained by protoplast transformation.Five targeted deletion mutants ?Fon00140,?Fon08535,?Fon01620,AFon08898 and ?Fon18919,and two cooresponding complement transfromants ?Fon00140-C and ?Fon08535-C were obtained.Mycelial growth phenotype,pathogenicity,stress response and extracellular hydrolytic enzyme were analyzed by comparing between the wild type,the targeted deletion mutants and complement transfromants.Main results are as follows:(1)No significant changes in mycelial growth,conidial production and germination,pathogenicity and other phenotypes of ?Fon01620,?Fon08898 and ?Fon18919 were observed,indicating that Fon01620,Fon08898 and Fon18919 are not involved in vegetative growth,conoidial production,and pathogenicity.The deletion mutants of?Fon01620,?Fon08898 and ?Fon18919 exhibited different phenotype in reponse to some stress response in general;however,all these deletion mutants showed increased sensitivity to cell wall perturbing agents,osmotic stress,oxidative stress and metal ion stress(e.g.CaC12)with exception of an increased resistance to metal ion stress factor MgCl2.(2)Functional analysis of Fon00140 and Fon08535 using deletion mutants revelaed that Fon00140 is not involved in mycelial growth,but is involved in the synthesis of pigments.Loss of Fon08535 affects the fungal development and pigment biosynthesis.The deletion mutants of both genes produced more conidia than the wild type but the morphology of the spores was affected as compared with the wild type.(3)The deletion mutants of Fon00140 and Fon08535 also showed different response to stress treatments.However,the deletion mutants showed an increased mycelial growth under osmotic stress(e.g.NaCl and Sorbitol).(4)The function of Fon00140 and Fon08535 in Fon pathogenicity was examined.The deletion mutants of Fon00140 and Fon08535 showed reduced penetration toward the root of watermelon seedlings.Similarly,in the deletion mutants,the expression levels of three improtant genes which were important in fusaric acid biosynthesis showed a significantly decrease.(5)The results revealed that the extracellular hydrolytic enzymes such as proteolytic enzyme,pectinase and cellulase were significantly reduced in the deletion mutants of Fon00140 and Fon08535.Results of this study revealed that Fon00140 and Fon08535 are important for various life processes of growth and development,environmental adaptability,pathogenicity and exocellular hydrolytic enzymes in F.oxysporum f.sp.nievum.