Investigating signaling components in the Rpg1-mediated disease resistance pathway

The barley stem rust-resistance gene Rpg1 confers resistance to many, but not all, pathotypes of the stem rust fungus Puccinia graminis f. sp. tritici. The Rpg1 gene was cloned by a map-based approach from barley cultivar Morex. Haplotype sequencing and transformation of the susceptible barley cultivar Golden Promise were employed to prove its identity. Rpg1 protein is a functionally active serine/threonine kinase with two tandem kinase domains.; To identify components involved in Rpg1-mediated disease resistance pathway, different approaches were performed. Mutational analysis in several plant-pathogen systems has been a powerful tool in identifying additional components that are required for R gene-mediated resistance. A fast-neutron induced rpr1 mutant, compromises Rpg1 -mediated resistance to P. graminis f. sp. tritici in the barley cv. Morex, which carries Rpg1. Therefore, the active form of deleted gene(s) is required for Rpg1 function. We have identified three candidate Rpr1 genes, Contig4901_s_at, HU03D17U_s_at, and Contig7061_s_at by transcript-based cloning using Barley1 GeneChip.; In order to further determine which candidate gene(s) is responsible for the rpr1 phenotype, we sequenced the deleted region encompassing HU03D17U_s_at and Contig4901_s_at. Sequence analysis of these 3 candidate genes based on computer predication and cDNA clones revealed possible gene structures. The third deleted gene, Contig7061_s_at, is predicted to be a functional serine/threonine protein kinase. Characterization of candidate Rpr1 genes provided basis for further investigation of the role of Rpr1 gene in Rpg1-mediated disease resistance.; To ascertain the global framework of host gene expression during early stage of rust fungus invasion, an mRNA expression analysis using Barley1 GeneChip was conducted in all pair-wise combinations of two isolines (Golden Promise and Rpg1 transgenic line) and two P. graminis f. sp. tritici pathotypes (MCC and QCC) across six time points after pathogen challenge. Our analysis showed that a total of 34 probe sets exhibited expression pattern differences between Golden Promise and Rpg1 transgenic line infected with Pgt-MCC, a total of 14 probe sets exhibited expression pattern differences between Pgt-MCC and Pgt-QCC inoculated onto Rpg1 transgenic line. These differentially expressed genes were activated correlating with fungal infection progression, suggesting that these early events might lead to resistance.