The Role of Nitric Oxide in Nitrogen Fixation by Legumes

Abstract

The legume-rhizobia symbiosis is an important process in agriculture because it allows the biological nitrogen fixation (BNF) which contributes to increasing the levels of nitrogen in the soil. Nitric oxide (⋅NO) is a small free radical molecule having diverse signaling roles in plants. Here we present and discuss evidence showing the role of ⋅NO during different stages of the legume-rhizobia interaction such as recognition, infection, nodule development, and nodule senescence. Although the mechanisms by which ⋅NO modulates this interaction are not fully understood, we discuss potential mechanisms including its interaction with cytokinin, auxin, and abscisic acid signaling pathways. In matures nodules, a more active metabolism of ⋅NO has been reported and both the plant and rhizobia participate in ⋅NO production and scavenging. Although ⋅NO has been shown to induce the expression of genes coding for NITROGENASE, controlling the levels of ⋅NO in mature nodules seems to be crucial as ⋅NO was shown to be a potent inhibitor of NITROGENASE activity, to induce nodule senescence, and reduce nitrogen assimilation. In this sense, LEGHEMOGLOBINS (Lbs) were shown to play an important role in the scavenging of ⋅NO and reactive nitrogen species (RNS), potentially more relevant in senescent nodules. Even though ⋅NO can reduce NITROGENASE activity, most reports have linked ⋅NO to positive effects on BNF. This can relate mainly to the regulation of the spatiotemporal distribution of ⋅NO which favors some effects over others. Another plausible explanation for this observation is that the negative effect of ⋅NO requires its direct interaction with NITROGENASE, whereas the positive effect of ⋅NO is related to its signaling function, which results in an amplifier effect. In the near future, it would be interesting to explore the role of environmental stress-induced ⋅NO in BNF.

Keywords: leghemoglobin; legumes; nitrogen fixation; reactive nitrogen species; reactive oxygen species; ⋅NO.

FIGURE 1

Nitric oxide participates at different stages of the nodulation process. ⋅NO was observed to be involved at all the different stages of nodulation, plant-rhizobium recognition (i), root hair curling (ii), infection thread (iii), nitrogen fixation in active nodules (iv), and senescing nodules (v). The light purple indicates the bacteria; the dark purple indicates the bacteroids; the gray indicates dead bacteroids; and the diffuse light green indicates ⋅NO. AA, amino acids; Phytogbs, PHYTOGLOBINS; Lbs, LEGHEMOGLOBINS; SR, systemic response.

FIGURE 2

The contribution of Phytogb/⋅NO cycle to ⋅NO production and NAD(P)H re-oxidation during hypoxia. ⋅NO is oxidized to nitrate by OXYPHYTOGLOBIN [Phytogb(Fe₂⁺)O₂], which turns to METPHYTOGLOBIN [Phytogb(Fe³⁺)]. Nitrate is then reduced to nitrite through NITRATE REDUCTASE (NR), and nitrite is reduced to ⋅NO though NR, the PLASMA MEMBRANE NITRITE-⋅NO REDUCATSE (NI-NOR) or the CYTOCHROME OXIDASE (COX). NAD(P)H oxidation occurs in the reactions forming ⋅NO and its subsequent oxidation back to nitrate.

FIGURE 3

Putative signaling effect of ⋅NO on nodule development. ⋅NO has been shown to induce the transcription of CRE1. Through this way, ⋅NO could promote CK signaling which is known to participate in nodule development. Moreover, ⋅NO is suggested to repress PIN expression resulting in auxin accumulation and cell division. Finally, the binding of ABA to the PYR/PYL/RCAR receptor can be avoided by tyrosine nitration, not allowing the inactivation of PP2C, which inhibits SnRK2 and the downstream ABA signaling through ABI5. ⋅NO is also known to directly inactivate both SnRK2 and ABI5 by S-nitrosylation, which also results in the suppression of ABA signaling. By these mechanisms, ⋅NO could attenuate ABA-mediated suppression of nodule development (shown in lower opacity). ABF1, ABA RESPONSIVE ELEMENT-BINDING FACTOR 1; ABI, ABA INSENSITIVE; CK, cytokinin; CRE1, CYTOKININ RESPONSE 1; NIN, NODULE INCEPTION; NSP2, NODULATION SIGNALING PATHWAY 2; PP2C, PROTEIN PHOSPHATASE OF THE TYPE IIC CLASS; SnRK2, SNF1-RELATED PROTEIN KINASE 2.

FIGURE 4

The effect of ⋅NO on nitrogen fixation and assimilation. (A) Effects of ⋅NO on NITROGENASE activity. (B) Detrimental effects of ⋅NO and RNS on nitrogen fixation and assimilation. In red are presented the interactions that lead to a lower nitrogen fixation and assimilation. COX, CYTOCHROME OXIDASE; GS, GLUTAMINE SYNTHASE.