ORA59 And EIN3 Interaction Couples Jasmonate-ethylene Synergistic Action To Antagonistic Salicylic Acid Regulation Of PDF 1.2 Expression

To survive the broad range of frequently encountered environmental perturbations,plants have developed sophisticated stress responses tailored to the nature of the stress and evolved delicate mechanisms for integration of endogenous and exogenous signals to adapt to the changing environment.Specifically,plants produce a blend of hormones,known as signal signature with varying quantities,compositions,and timing depending on the lifestyle and the nature of the stress.In addition to the signal signature,plants also rearrange various hormone-signaling pathways to effectively counter these challenges while maintaining a fine balance for resources simultaneously required for growth and development.The plant hormone Salicylic Acid(SA)is a phenolic compound that plays a key role in plant defense responses primarily against biotrophic pathogens.Activation of the SA pathway at the site of infection triggers long distance resistance known as systemic acquired resistance(SAR)to prevent new infections and to limit the spread of the already existing infection.Jasmonic Acid(JA)and its structurally related metabolites wild typelectively known as jasmonates arc oxygenated lipid-derived metabolites produced rapidly by the oxylipin biosynthesis pathway in response to environmental and devvelopmental signals.The JA signaling pathway consists of two branches;the MYC branch coupled to wounding and defense against insect herbivores,and the ERF branch that is associated with enhanced resistance to nccrotrophic pathogens.Hormonal crosstalk is central for tailoring plant responses to the nature of challenges encountered.The regulation of antagonistic and synergistic interplay between salicylic acid and jasmonic acid is one of the key strategies plants deploy to organize effective stress response signaling networks.Superimposed on this,there are interactions with other hormones such as ethylene(ET),that further verify the complexity and significance of hormonal interaction in maximizing plants' ability to effectively mount a well-designed adaptive response against a specific stress.The role of antagonism between the two major defense hormones,salicylic acid and jasmonic acid,and modulation of this interplay by ethylene in favor of JA signaling pathway in plant stress responses is well recognized,but the underlying mechanism is not fully understood.Accumulation of PDF 1.2 mRNA is commonly used as a marker for activation of the JA/ET signaling pathway.To understand what this marker actually reports it is crucial to understand the regulation of PDF 1.2 mRNA accumulation.In Arabidopsis,the SA-mediated suppression and ET/JA-induction of PDF 1.2 establish this marker of the ERF branch of the JA pathway as an ideal target for examining the underlying mechanism of the hormonal crosstalk regulating effective and tailored stress responses.Our research found:1.EIN 3/EIL 1 is required for SA-suppression and JA-induction of PDF 1.2 expression.For these experiments,we specifically compared the data obtained both from soil and plate grown Wild type plants,and show JA-mediated induction and SA-mediated suppression of PDF 1.2 expression levels are more pronounced in soil,a condition closer to the natural conditions.In soil-grown plants,exogenous application of either JA,SA and JA+SA strongly induces PDF 1.2 expression,presumably because the plants already produce low amounts of the signaling molecules.2.The highly induced PDF 1.2 in response to SA/JA combined application imply synergistic/additive roles of these hormones,and additionally display a molecular link between their crosstalk via ORA 59 and the EIN 3/EIL 1 transcription factors.3.SA treatment as compared to mock resulted to markedly increased levels of the ERF 1 transcript levels both in the Wild type and ein 3/eil 1 mutant lines,albeit partially EIN 3/EIL 1 dependent and most notably in Wild type.4.SA-mediated reduced stability of ORA 59 protein is dependent on EIN 3/EIL 1,but the JA-induced enhanced stability of ORA 59 is not.5.EIN 3 and ORA 59 are nuclear co-localized and physically interact.Identified the auto activation domain of ORA 59,and further confirmed protein-protein interaction between EIN 3 and ORA 59 by using Y2H.6.EIN 3 mediated regulation of the protein and not the transcript levels of ORA 59.Specifically,EIN 3 binds to and mediates targeting of ORA 59 for degradation by the 26S proteasome machinery.7.Exogenous application of SA results in increased EIN 3 protein abundance,and affects the ethylene-signaling pathway.