Functional Analysis Of Eda,A Crucial Gene To Pathogenicity Of Pectobacterium Carotovorum Subsp. Carotovorum

Pectobacterium carotovorum subsp.carotovorum?Pcc?is an important plant pathogen causeing soft rot disease.The main pathogenic factor of Pccisplant cell wall degrading enzymes?PCWDEs?including pectinase,protease and cellulase,among these PCWDEs,pectinase is the most important one.Soft rot disease caused by Pcc is worldwide in many crops and ornamential plants resulting in serious economical loss.The understanding about the pathogenicity clearly will be of benefit sfor setting strategies for soft rot disease control.Previous studies in our laboratory found that glucose-6-phosphate dehydrogenase?Zwf,PC 1 1832?expressed up regulate when PccS 1 interacted with Zantedeschia elliotiana in vivo.The results from the assays of pathogenicity showed that the virulence of the gene insertional mutant of zwf single-crossoveredin homologous recombination was significantly reduced,while that of zwf gene deletion mutantwas remained,comparing with the wild type strain of PccSl,the deletion mutant of eda?2-keto-3-deoxy-6-phosphogluconate aldolase gene,PC1₁1833?,a gene downstream of zwf in the same direction,showed a significant attenuated in pathogenicity on host and the activities of pectinase,and lost the survival ability on the media of sodium acetate as a sole carbon source.Eda,KdgK?2-dehydro-3-deoxygluconokinase,coding by kdgK,upstream of eda?and Zwf are all the key enzymesin intracellular pectin degradation and Entner-Doudoroff pathway?ED pathway?.In this work,it was oberserved that the pathogenicity of the insertional mutant of zwf single-crossoveredhad been recovered by complement with eda but not with zwf itself,there existed a couple of transcription between zwf and eda.Meanwhile,the kdgK deletion mutant showed no obvious changes on the pathogenicity just like the mutant of zwf knocked out.On the other hand,ED pathway could be recovered by complement of edd from Pectobacerium atrosepticum,and H+ translocation ability of the complementary strain was improved,but the pathogenicity exhibited novariation comparing with the wild type.Glyoxylate cycle is nessessary for carbonic circulation and energy metabolismwhen acetic acid is the sole carbon source,which includes three key enzymes,isocitrate dehydrogenase kinase?AceK,encode by PC1₃777?,isocitrate lyase?AceA,by PC 3778?and malate synthase A?AceB,by PC1₃779?,mutants of each of the three genes deletedwere all abolished survival ability on C2 as sole carbon source as ? eda did,but only ? acaK showed virulence no the host.It was also found that the expression of these three genes were all siginificantly attenuated in ? eda.Furthermore,Eda showed interaction with itself in Co-IP and the yeast two-hybrid verification.So,we speculate that functions of eda on pathgenicity is not through carbon flux of the metabolic pathways in which eda takes part in,including ED pathway and pectin degradation,indeed through the regulations of Eda in some mechanisms undiscovered,and that eda affects the expression of genes coding enzymes for glyoxylate cycle,and it might function in a pattern of polymeride.