Both plant and bacterial nitrate reductases contribute to nitric oxide production in Medicago truncatula nitrogen-fixing nodules.
Nitric oxide (NO) is a signaling and defense molecule of major importance in living organisms. In the model legume Medicago truncatula, NO production has been detected in the nitrogen fixation zone of the nodule, but the systems responsible for its synthesis are yet unknown and its role in symbiosis is far from being elucidated. In this work, using pharmacological and genetic approaches, we explored the enzymatic source of NO production in M. truncatula-Sinorhizobium meliloti nodules under normoxic and hypoxic conditions. When transferred from normoxia to hypoxia, nodule NO production was rapidly increased, indicating that NO production capacity is present in functioning nodules and may be promptly up-regulated in response to decreased oxygen availability. Contrary to roots and leaves, nodule NO production was stimulated by nitrate and nitrite and inhibited by tungstate, a nitrate reductase inhibitor. Nodules obtained with either plant nitrate reductase RNA interference double knockdown (MtNR1/2) or bacterial nitrate reductase-deficient (napA) and nitrite reductase-deficient (nirK) mutants, or both, exhibited reduced nitrate or nitrite reductase activities and NO production levels. Moreover, NO production in nodules was found to be inhibited by electron transfer chain inhibitors, and nodule energy state (ATP-ADP ratio) was significantly reduced when nodules were incubated in the presence of tungstate. Our data indicate that both plant and bacterial nitrate reductase and electron transfer chains are involved in NO synthesis. We propose the existence of a nitrate-NO respiration process in nodules that could play a role in the maintenance of the energy status required for nitrogen fixation under oxygen-limiting conditions.
UMR INRA 1301, CNRS 6243, Université Nice Sophia Antipolis, Interactions Biotiques et Santé Végétale, Institut Agrobiotech, 06903 Sophia Antipolis cedex, France.
Faouzi
F
Horchani
Marianne
M
Prévot
Alexandre
A
Boscari
Edouard
E
Evangelisti
Eliane
E
Meilhoc
Claude
C
Bruand
Philippe
P
Raymond
Eric
E
Boncompagni
Samira
S
Aschi-Smiti
Alain
A
Puppo
Renaud
R
Brouquisse
D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D015687:Cell Hypoxia; D015964:Gene Expression Regulation, Bacterial; D018506:Gene Expression Regulation, Plant; D055785:Gene Knockdown Techniques; D046913:Medicago truncatula / Q000201:enzymology* / Q000235:genetics / Q000382:microbiology; D008928:Mitochondria / Q000201:enzymology; D009565:Nitrate Reductases / Q000235:genetics / Q000378:metabolism*; D009566:Nitrates / Q000494:pharmacology; D009569:Nitric Oxide / Q000096:biosynthesis*; D009573:Nitrites / Q000494:pharmacology; D009586:Nitrogen Fixation; D010100:Oxygen / Q000502:physiology; D010940:Plant Proteins / Q000235:genetics / Q000378:metabolism; D034622:RNA Interference; D053204:Root Nodules, Plant / Q000201:enzymology / Q000382:microbiology / Q000502:physiology*; D016962:Sinorhizobium meliloti / Q000201:enzymology* / Q000235:genetics / Q000502:physiology; D013559:Symbiosis; D017973:Tungsten Compounds / Q000494:pharmacology
D001426:Bacterial Proteins; D009566:Nitrates; D009573:Nitrites; D010940:Plant Proteins; D017973:Tungsten Compounds; D009569:Nitric Oxide; D009565:Nitrate Reductases; D010100:Oxygen; C045951:tungstate