Identification And Functional Analysis Of The Targets Of Effector Protein Pst06941 In Puccinia Striiformis F.sp.Tritici

Wheat stripe rust,caused by the obligate biotrophic fungus Puccinia striiformis f.sp.tritici?Pst?,often imposes a tremendous threat to the wheat production worldwide.As a result of constant pathogenicity variation,the resistance of the existing wheat cultivars has weakened or even lost.Therefore,studying the pathogenic mechanism of virulence factor from Pst,especially the pathogenic mechanism of effector,has important theoretical basis for controlling stripe rust.In the previous study,Pst06941 has been confirmed to be a typical effector that contributes to the pathogenicity of Pst via bioinformatics analysis,the yeast secretion system assays,agrobacterium-mediated transient expression system and virus induced gene silencing technology?VIGS?.In our study,the interaction between Pst06941 and TaNADH dehydrogenase subunit I?TaNADH ??was confirmed by yeast two-hybrid assay,Bimolecular Fluorescence Complementation and Pull-down assays.In plants,NADH dehydrogenase is usually ubiquitous in the form of a complex.NADH I is a complex that plays an important role in the process of electron transfer.As the main entrance to transfer chain for the outside electronics,NADH I also functions as a proton pump.In order to further clarify the pathogenesis of Pst06941,we used bioinformatics analysis and VIGS technology to explore the function of the target TaNADH ?.Sequence analysis showed that the ORF of TaNADH ? gene consists of 181 amino acid residues and contains only one evolutionarily conserved Fe-S protein domain.qRT-PCR assays revealed that TaNADH ? is up-regulated during early infection stages of Pst.Silencing of TaNADH ? in wheat by VIGS technique significantly reduced wheat resistance to Pst.The cytological observation revealed that the infection area was increased at 24 hpi and 120 hpi.The accumulation area of H₂O₂ induced by the incompatible Pst strain CYR23 was down-regulated at 24,48 and 72 hpi,and the expression levels of TaPR1 and TaPR2 were also decreased.Overall,our data indicate that Pst06941 targets TaNADH dehydrogenase subunit I related to host immune responses during Pst infection,suggesting that Pst06941may affect host immune responses in which TaNADH dehydrogenase subunit I participates to promote Pst infection.