WKS1-based Proteomics And Molecular Recombination Of WKS Genes For Strip Rust Resistance In Wheat

Wheat stripe rust, caused by Puccinia striiformis f. sp. tritici (Pst), is one of the mostdevastating diseases worldwide.The resistant gene of WKS1which was cloned from Triticumturgidum ssp. dicoccoides offers a broad spectrum resistance to many Pst races. Compared toother resistant genes, it contains two domains, kinase and START. Thefore, the Yr36gene isnamed Wheat Kinase-START1(WKS1). It can effectively reduce stripe rust demage, and cutthe production loss. WKS1has a close homologue Wheat Kinase-START2(WKS2). The twohomologues share84.5%similarity in protein levels. However, the molecular mechanism ofWKS1gene and the function of WKS2gene are largely unknown. In this study, the followingexperiments were performed to increase our understanding on WKS genes.(1) WKS1-based proteomics for wheat stripe rust resistance: The study was performedon the Yr36isogenic lines, LDN without WKS1gene and RSL65with Yr36gene, using theiTRAQ（isobaric tags for relative and absolute quantitation）technology. Four differentsamples, RSL65(rust infected tissue, In), RSL65(control, CK), LDN (In), and LDN (CK),were investigated to identify differentially expressed proteins relating to the WKS1gene. Intotal,861proteins were identified. In comparison to LDN (CK), RSL65(In), RSL65(CK), andLDN (In) shared58differentially expressed proteins, of which26proteins were up regulatedand32proteins were down regulated. Most of differentially expressed proteins are related toorganelle composion, photosynthesis, binding, and transcriptional activation. They are likelyinvolved in metabolism, signal transduction, defense and stress responses.(2) Molecular design of WKS genes: We cloned six WKS cDNA homologues (WKS1orWKS2）from Aegilops comosa, Thinopyrum elongatum, Th. bessarabicum, and Triticumturgidum ssp. dicoccoides. Eight recombinant genes were created by motif swapping betweenKinase and START. All native and derived WKS genes were used to construct Ubi::WKSexpression vectors. To validate the genes function, they were transformed into arust-susceptible accession of Brachypodium distachyon. Stable transgenic events and cDNAtranscription of investigated WKS genes were confirmed in T0generation. The study lays afoundation for introducing novel WKS genes and revealing molecular mechanism of WKS1function.