title: Production of nitric oxide and nitrosylleghemoglobin complexes in soybean nodules in response to flooding.
abstract: Nitric oxide (NO) has gained interest as a major signaling molecule during plant development and in response to environmental cues. Formation of NO during symbiotic interactions has been reported, but the role and sources of NO in nodules remain unclear. In this work, the involvement of denitrification, performed by the symbiont Bradyrhizobium japonicum, in NO formation in soybean nodules in response to flooding conditions has been investigated by inoculating plants with napA-, nirK-, or norC-deficient mutants. Levels of nitrosylleghemoglobin (LbNO) in flooded nirK and norC nodules were significantly higher than those observed in wild-type nodules. In addition, nirK and norC nodules accumulated more nitrite and NO, respectively, than wild-type nodules. By contrast, levels of LbNO, nitrite, and NO in flooded napA nodules were lower than in wild-type nodules. These results suggest that LbNO formation in soybean nodules in response to flooding conditions is caused by nitrite and NO generated from periplasmic nitrate reductase (Nap) and also containing nitrite reductase (NirK) denitrification enzymes. Flooding caused a decrease of nifH expression and nitrogenase activity in wild-type and norC nodules but not in napA or nirK nodules. Incubation of wild-type and norC nodules with a NO scavenger counteracted the effect of flooding. Under free-living conditions, beta-galactosidase activity from a nifD'-'lacZ fusion decreased in a norC mutant, which also accumulated NO in the medium. These results suggest that NO formed by Cu-containing nitrite reductase in soybean nodules in response to flooding has a negative effect on expression of nitrogenase. We propose that Lb has a major role in detoxifying NO and nitrite produced by bacteroidal denitrification in response to flooding conditions.
authors.affiliation: Estación Experimental del Zaidín, Granada, Spain.
authors.forename: Cristina
authors.initials: C
authors.lastname: Sánchez
authors.forename: Andrew J
authors.initials: AJ
authors.lastname: Gates
authors.forename: Georgina E
authors.initials: GE
authors.lastname: Meakin
authors.forename: Toshiki
authors.initials: T
authors.lastname: Uchiumi
authors.forename: Lourdes
authors.initials: L
authors.lastname: Girard
authors.forename: David J
authors.initials: DJ
authors.lastname: Richardson
authors.forename: Eulogio J
authors.initials: EJ
authors.lastname: Bedmar
authors.forename: María J
authors.initials: MJ
authors.lastname: Delgado
mesh_terms: D000114:Acetylene / Q000378:metabolism; D020369:Bradyrhizobium / Q000187:drug effects / Q000201:enzymology / Q000235:genetics; D004578:Electron Spin Resonance Spectroscopy; D055868:Floods; D015964:Gene Expression Regulation, Bacterial / Q000187:drug effects; D018506:Gene Expression Regulation, Plant / Q000187:drug effects; D005798:Genes, Bacterial / Q000235:genetics; D007874:Leghemoglobin / Q000096:biosynthesis*; D009154:Mutation / Q000235:genetics; D009569:Nitric Oxide / Q000096:biosynthesis*; D009591:Nitrogenase / Q000235:genetics / Q000378:metabolism; D010084:Oxidation-Reduction / Q000187:drug effects; D010269:Paraquat / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D053204:Root Nodules, Plant / Q000187:drug effects / Q000235:genetics / Q000378:metabolism* / Q000382:microbiology; D013025:Glycine max / Q000187:drug effects / Q000235:genetics / Q000378:metabolism* / Q000382:microbiology; D014867:Water / Q000494:pharmacology*; D001616:beta-Galactosidase / Q000378:metabolism
chemical_list: D007874:Leghemoglobin; D014867:Water; D009569:Nitric Oxide; D009591:Nitrogenase; D001616:beta-Galactosidase; D000114:Acetylene; D010269:Paraquat