ASSESSING NITRIC OXIDE IN PLANT DEFENSE USING NO SPIN TRAPPING

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This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. NO acts as a signaling molecule in a wide range of physiological processes in both eukaryotes and prokaryotes. In plants, NO participates in growth, development, flowering, and response to biotic and abiotic stresses. It has been shown that nitric oxide plays an important role in plant disease resistance. However, unlike our appreciation of NO functioning in animals, the origin and the quantification of plant NO responses to stimuli remain unclear. A comprehensive understanding of NO biology is dependent on the development of reliable NO detection methods, and EPR represents a reliable tool in this respect. Stabilization of NO radical with spin trap complexes and its detection by ESR offer the advantage of being both selective and sensitive. Initial ESR study in ACERT using NO spin trap Fe(DETC)2 showed that the model plant Arabidopsis thaliana responds to the stress caused by infiltration with Pseudomonas syringae by more rapid and robust production of NO. The Dan Klessig group is interested in using lipophilic Fe(DETC)2 complex or hydrophilic Fe(MGD)2 spin traps to evaluate NO formation in the context of plant-microbe interactions in 1) plant cell suspensions (tobacco cells) elicited for NO production with cryptogein, a 10 kDa protein secreted by the oomycete Phytophtora cryptogea that induces host defense responses, and 2) in Arabidopsis plants infected with either a bacterial pathogen (Pseudomonas syringae) or a peptide elicitor corresponding to the conserved domain of bacterial flagellin. We currently use 15N substitution to exploit the ability of the ESR measurement to distinguish between 14NO and 15NO, and we are confident that an ESR-based approach will help to elucidate the origin(s) of this radical in planta.