Molecular responses of legumes to abiotic stress: post-translational modifications of proteins and redox signaling

Abstract

Legumes include several major crops that can fix atmospheric nitrogen in symbiotic root nodules, thus reducing the demand for nitrogen fertilizers and contributing to sustainable agriculture. Global change models predict increases in temperature and extreme weather conditions. This scenario might increase plant exposure to abiotic stresses and negatively affect crop production. Regulation of whole plant physiology and nitrogen fixation in legumes during abiotic stress is complex, and only a few mechanisms have been elucidated. Reactive oxygen species (ROS), reactive nitrogen species (RNS), and reactive sulfur species (RSS) are key players in the acclimation and stress tolerance mechanisms of plants. However, the specific redox-dependent signaling pathways are far from understood. One mechanism by which ROS, RNS, and RSS fulfil their signaling role is the post-translational modification (PTM) of proteins. Redox-based PTMs occur in the cysteine thiol group (oxidation, S-nitrosylation, S-glutathionylation, persulfidation), and also in methionine (oxidation), tyrosine (nitration), and lysine and arginine (carbonylation/glycation) residues. Unraveling PTM patterns under different types of stress and establishing the functional implications may give insight into the underlying mechanisms by which the plant and nodule respond to adverse conditions. Here, we review current knowledge on redox-based PTMs and their possible consequences in legume and nodule biology.

Keywords: Abiotic stress; legumes; nitric oxide; nitrogen fixation; post-translational modifications; reactive oxygen/nitrogen/sulfur species; redox signaling; symbiosis.

Fig. 1.

Scheme showing the infection process, the major differences between indeterminate and determinate nodules, and some key metabolic pathways in the nodule cells. (A) Briefly, the infection process is as follows: legume roots release flavonoids to the rhizosphere which induce the production of nodulation (Nod) factors in compatible rhizobia (depicted as red dots next to root hairs); Nod factors are recognized by root receptors that activate the symbiotic signaling pathway; rhizobia enter the root through hairs that curve and trap the bacteria inside a curl; invaginations of the cell membrane form infection threads that permit the invasion of the root cortex by rhizobia; a new nodule meristem forms underneath the site of infection; as the nodule grows, the bacteria are released into membrane-bound compartments, the symbiosomes, inside the nodule cells, where the bacteria differentiate into bacteroids and start N₂ fixation. As a result, two major types of nodules are formed. Indeterminate nodules of Medicago truncatula and crops such as pea and alfalfa contain a persistent meristem and are generally elongated with a longitudinal gradient of age. Four zones can be distinguished from the apex (distal) to the base (proximal) regions: zone I (meristem), zone II (infection), zone III (N₂-fixing), and zone IV (senescent). Determinate nodules of Lotus japonicus and of crops such as soybean and common bean lack permanent meristems and are usually spherical. In this case, N₂ fixation takes place in the central infected zone, which also contains uninfected or interstitial cells. Abbreviations of cell layers: c, cortex; e, epidermis; en, endodermis; p, pericycle; vb, vascular bundle. (B) Some key processes in the symbiotic nodule cells. Sucrose from the shoot is metabolized to malate that is transported into bacteroids through dicarboxylate transporters (Dct). In the bacteroids, malate is oxidized, providing energy through the tricarboxylic acid (TCA) cycle and the electron transport chain (ETC) for nitrogenase (N₂ase) activity. Fixed nitrogen in the form of ammonium is transported back to the plant where, with some exceptions, it is assimilated into the amides glutamine and asparagine in indeterminate nodules or ureides in determinate nodules. In the cytoplasm of infected cells, leghemoglobin (Lb) transports and delivers O₂ to the symbiosomes at a low steady concentration to avoid the inhibition of nitrogenase, but to simultaneously allow high rates of bacteroid respiration. After Oldroyd et al., (2011), Popp and Ott (2011), and Hichri et al., (2015).

Fig. 2.

Redox-dependent PTMs. Met residues can be oxidized by hydrogen peroxide (H₂O₂) to Met sulfoxides. The S and R stereoisomers are specifically reduced back to Met by methionine sulfoxide reductases A and B, respectively. Oxidation of deprotonated thiols of Cys residues (–S^-) by H₂O₂ leads to the formation of sulfenic acid (–SOH), which may react with another thiol to form disulfides (–S–S–). This modification can be reverted by thioredoxins and glutaredoxins. The –SOH group can be an intermediate to other redox modifications (see below) or be further oxidized to sulfinic acid (–SO₂H) and sulfonic acid (–SO₃H). S-nitrosylation (–SNO) is mostly mediated by nitrogen oxides and trans-nitrosylating agents such as S-nitrosoglutathione (GSNO). S-glutathionylation (–SSG) occurs by two main mechanisms: reaction of the target protein with GSNO, and reaction of reduced glutathione (GSH) with –SOH. The reaction of hydrogen sulfide (HS^-) with –SOH, –SNO, –SSG, or disulfide bridges induces persulfidation (–SSH). Peroxynitrite (ONOO^-) is formed by the reaction of nitric oxide (NO) with superoxide (O₂^.-) radicals. In turn, radicals derived from ONOO^- breakdown oxidize Tyr residues to tyrosyl radicals; these react with nitrogen dioxide (NO₂), produced from ONOO^- decomposition, to yield NO₂–Tyr. The direct oxidation of Lys, Arg, Pro, and Thr by hydroxyl radicals (·OH) incorporates the carbonyl moiety into proteins. Alternatively, oxidation of a polyunsaturated fatty acid (PUFA; a simplified representation is shown lacking part of the aliphatic chain) produces unstable lipid hydroperoxides that decompose to secondary products known as reactive carbonyl species (RCSs). These react with amino acid side chains and generate carbonyl derivatives. Moreover, Arg and Lys residues may react with reducing sugars or α-dicarbonyls such as glyoxal and methylglyoxal, generating glycation products that are readily oxidized to form relatively stable advanced glycation end products (AGEs).

Fig. 3.

Expression profile of genes involved in redox homeostasis in the shoots of Lotus japonicus plants exposed to drought or salt stress. Gene up-regulation (>two-fold) and down-regulation (<0.5-fold) are indicated in red and blue, respectively. Gene IDs are given according to the L. japonicus MG-20 genome v3.0 and data were retrieved from the L. japonicus Expression Atlas (Lotus Base; https://lotus.au.dk). Transcriptomic data under salt and drought stress were published, respectively, by Sanchez et al., (2008) and Díaz et al., (2010). Abbreviations: ALDH, aldehyde dehydrogenase; AR, alkenal reductase; DR, dehydroascorbate reductase; Glb, phytoglobin; Grx, glutaredoxin; GSHS, glutathione synthetase; GSTU, glutathione transferase tau family; Msr, methionine sulfoxide reductase.

Fig. 4.

Expression profile of genes involved in redox homeostasis in the roots and shoots of Medicago truncatula plants exposed to drought or salt stress. Gene up-regulation (>two-fold) and down-regulation (<0.5-fold) are indicated in red and blue, respectively. Gene IDs are given according to the M. truncatula genome v4.0. Transcriptomic data of roots and shoots under drought stress were retrieved from the Gene Expression Atlas (https://mtgea.noble.org/v3/) and those of salt stress in the roots were published by Li et al., (2009). Gene sequences were obtained from Legume IP (http://plantgrn.noble.org/LegumeIP/gdp/), except for Gpx1-2 and GSTU19, which were obtained from GenBank. The probes used to determine gene expression profiles are listed in Table 1. Abbreviations: AKR, aldo-keto reductase; ALDH, aldehyde dehydrogenase; Apx, ascorbate peroxidase; AOR, alkenal/one oxidoreductase; DR, dehydroascorbate reductase; γECS, γ-glutamylcysteine synthetase; GalDH, L-galactono-1,4-lactone dehydrogenase; GGP, GDP-L-galactose phosphorylase; Glb, phytoglobin; GLX, glyoxalase; Gpx, glutathione peroxidase; GR, glutathione reductase; Grx, glutaredoxin; GSHS, glutathione synthetase; GSNOR, S-nitrosoglutathione reductase; GSTU, glutathione S-transferase tau family; hGSHS, homoglutathione synthetase; Lb, leghemoglobin; MR, monodehydroascorbate reductase; Msr, methionine sulfoxide reductase; Prx, peroxiredoxin; SOD, superoxide dismutase; Trx, thioredoxin.

Fig. 5.

Expression profiles of genes involved in redox homeostasis in nodules of Lotus japonicus and Medicago truncatula following two or four days of drought stress. Gene up-regulation (>two-fold) and down-regulation (<0.5-fold) are indicated in red and blue, respectively. Expression data were retrieved from LegumeIP v3 (http://plantgrn.noble.org/LegumeIP/gdp/) and Sańko-Sawczenko et al., (2019). Abbreviations: ALDH, aldehyde dehydrogenase; Glb, phytoglobin; Grx, glutaredoxin; GSTU, glutathione transferase tau family; Lb, leghemoglobin; Prx, peroxiredoxin; SOD, superoxide dismutase; Trx, thioredoxin.