Dissecting disease resistance mechanisms using Arabidopsis dnd 'defense, no death' mutants

The defense, no death (dnd) mutants of Arabidopsis thaliana were previously identified in a genetic screen for lines that failed to activate the hypersensitive response (HR) when inoculated with Pseudomonas syringae expressing avrRpt2. The dnd1 and dnd2 mutants exhibit enhanced disease resistance to virulent and avirulent P. syringae , high levels of salicylic acid (SA), and constitutive pathogenesis-related (PR) gene expression despite loss of the HR. The dnd1 phenotypes were due to the mutation of AtCNGC2, which encodes a cyclic nucleotide-gated (CNG) ion channel. In one segment of the present study, I discovered that DND2 encodes another CNG ion channel, AtCNGC4.; The dnd mutants and other Arabidopsis mutants are useful genetic tools to decipher the mechanisms that regulate defense signaling and the HR. In the present study, mutant alleles of NPR1, SID2, NDR1, and EIN2 were introduced into the dnd mutant backgrounds. Defense phenotypes such as resistance to P. syringae, Peronospora parasitica, and Botrytis cinerea as well as PR gene expression and the HR were examined in dnd double and triple mutants. Mutant alleles of sid2 and ndr1 partially relieved dnd1-mediated resistance to P. syringae expressing avrRpt2, whereas npr1 had no detectable effect on resistance to virulent or avirulent P. syringae. The resistance of dnd1 plants to P. parasitica was compromised by npr1, sid2, ndr1, but not ein2. Conversely, dnd1 ein2 and dnd1 npr1 ein2 plants exhibited enhanced susceptibility to B. cinerea, while mutants with defects in SA-mediated signaling did not alter dnd1-mediated resistance. At the molecular level, npr1 and sid2 mutations reduced/eliminated constitutive PR-1 gene expression of dnd1 plants, but intriguingly, caused elevation of PDF1.2 expression indicating potential crosstalk between SA-mediated and JA/ethylene-mediated signaling pathways. Results from RNA analysis and GUS staining of dnd1 npr1 plants suggest that dnd1 plants can activate PR gene pathways in an NPR1-independent fashion. It was further demonstrated that the HR- phenotype displayed by dnd1 can be overcome by introducing ndr1 or sid2 mutant alleles. Genetic dissection of the dnd mutants has provided greater insight regarding the interrelationships among defense signaling networks.