Old poisons, new signaling molecules: the case of hydrogen cyanide.

The high phenotypic plasticity developed by plants includes rapid responses and adaptations to aggressive or changing environments. To achieve this, they evolved extremely efficient mechanisms of signaling mediated by a wide range of molecules, including small signal molecules. Among them, hydrogen cyanide (HCN) has been largely ignored due to its toxic characteristics. However, not only is it present in living organisms, but it has been shown that it serves several functions in all kingdoms of life. Research using model plants has changed the traditional point of view, and it has been demonstrated that HCN plays a positive role in the plant response to pathogens independently of its toxicity. Indeed, HCN induces a response aimed at protecting the plant from pathogen attack, and the HCN is provided either exogenously (in vitro or by some cyanogenic bacteria species present in the rhizosphere) or endogenously (in reactions involving ethylene, camalexin, or other cyanide-containing compounds). The contribution of different mechanisms to HCN function, including a new post-translational modification of cysteines in proteins, namely S-cyanylation, is discussed here. This work opens up an expanding 'HCN field' of research related to plants and other organisms.

Avoidant/resistant rather than tolerant olive rootstocks are more effective in controlling Verticillium wilt.

The identification of rootstocks of low susceptibility to Verticillium dahliae can become a valuable procedure to achieve effective control of Verticillium wilt in the olive grove. This not only involves the identification of suitable genotypes, but also the study of the interaction between the rootstock and the grafted scion. Thus, a rootstock that prevents or minimizes V. dahliae proliferation (avoidance/resistance strategy) can have very different effects on a susceptible scion compared to a rootstock that shows few or no symptoms despite being infected (tolerance strategy). Both resistance and tolerance mechanisms have been recently identified in wild olive genotypes with low susceptibility to V. dahliae. When used as rootstocks of the highly susceptible variety 'Picual', we found that resistant genotypes, including the cultivar 'Frantoio', were more effective than tolerant genotypes in controlling Verticillium wilt. Furthermore, tolerant genotypes were as ineffective as susceptible or extremely susceptible genotypes in controlling Verticillium wilt. We also identified rootstock-scion combinations with behaviours that were not expected according to the degree of susceptibility previously observed in the non-grafted rootstock. Although the rootstocks were able to control Verticillium wilt according to its degree of susceptibility to V. dahliae, the ability to control the infection was not adequately transferred to the grafted scion. Our results confirmed that: the degree of susceptibility to Verticillium wilt of an olive variety does not predict its performance as a rootstock; to use a very low susceptible genotype as rootstock of a susceptible scion increases the susceptibility of the genotype used as rootstock; in any case, avoidant/resistant rootstocks are more effective than tolerant rootstocks in reducing the susceptibility of the grafted plant to V. dahliae.

Growth Quality and Development of Olive Plants Cultured In-Vitro under Different Illumination Regimes.

Light-emitting diodes (LEDs) are useful for the in-vitro micropropagation of plants, but little information is available on woody species. This work compares the effects of light quality and intensity on the growth and development of micropropagated olive plants from two different subspecies. Illumination was provided with fluorescent and LED lamps covering different red/blue ratios (90/10, 80/20, 70/30, 60/40) or red/blue/white combinations, as well as different light intensities (30, 34, 40, 52, 56, 84, 98 and 137 µmol m-2 s-1 of photosynthetic photon fluxes, PPF). Olive plants exhibited high sensitivity to light quality and intensity. Higher red/blue ratios or lower light intensities stimulated plant growth and biomass mainly as a consequence of a higher internodal elongation rate, not affecting either the total number of nodes or shoots. In comparison to fluorescent illumination, LED lighting improved leaf area and biomass, which additionally was positively correlated with light intensity. Stomatal frequency was positively, and pigments content negatively, correlated with light intensity, while no clear correlation was observed with light quality. In comparison with fluorescent lamps, LED illumination (particularly the 70/30 red/blue ratio with 34 µmol m-2 s-1 PPF intensity) allowed optimal manipulation and improved the quality of in-vitro micropropagated olive plants.

Wild Olive Genotypes as a Valuable Source of Resistance to Defoliating Verticillium dahliae.

Resistance to the defoliating pathotype of Verticillium dahliae has been evaluated in a pool of 68 wild genotypes of olive belonging to the SILVOLIVE collection. Resistance was evaluated by assessing symptom severity using a 0-4 rating scale, estimating the relative area under the disease progress curve (RAUDPC), determining the percentage of dead plants (PDP), and measuring the evolution of morphological parameters in inoculated plants over time. In addition, the density levels of V. dahliae in the stem of root-inoculated genotypes have been quantified by means of quantitative real-time PCR at 35 and 120 days after inoculation (dai). Fifteen genotypes (22%) were cataloged as resistant to V. dahliae (i.e., disease parameters did not significantly differ from those of the resistant cultivar Frantoio, or were even lower). Resistant genotypes are characterized by presenting fewer symptoms and a lower amount of V. dahliae DNA at 120 dai than at 35 dai, indicating their ability to control the disease and reduce the density of the pathogen. The rest of the evaluated genotypes showed variable levels of susceptibility. Overall analysis of all genotypes showed high correlation between symptomatology and the amount of V. dahliae DNA in the stem of inoculated genotypes at 120 dai, rather than at 35 dai. However, correlation at 120 dai was not observed in the set of resistant genotypes, suggesting that resistance to defoliating V. dahliae in olive is based on the occurrence of different mechanisms such as avoidance or tolerance. These mechanisms are valuable for designing breeding programs and for the identification of target genes and resistant rootstocks to better control Verticillium wilt in the olive grove.

Chloride nutrition improves drought resistance by enhancing water deficit avoidance and tolerance mechanisms.

Chloride (Cl-), traditionally considered harmful for agriculture, has recently been defined as a beneficial macronutrient with specific roles that result in more efficient use of water (WUE), nitrogen (NUE), and CO2 in well-watered plants. When supplied in a beneficial range of 1-5 mM, Cl- increases leaf cell size, improves leaf osmoregulation, and reduces water consumption without impairing photosynthetic efficiency, resulting in overall higher WUE. Thus, adequate management of Cl- nutrition arises as a potential strategy to increase the ability of plants to withstand water deficit. To study the relationship between Cl- nutrition and drought resistance, tobacco plants treated with 0.5-5 mM Cl- salts were subjected to sustained water deficit (WD; 60% field capacity) and water deprivation/rehydration treatments, in comparison with plants treated with equivalent concentrations of nitrate, sulfate, and phosphate salts. The results showed that Cl- application reduced stress symptoms and improved plant growth during water deficit. Drought resistance promoted by Cl- nutrition resulted from the simultaneous occurrence of water deficit avoidance and tolerance mechanisms, which improved leaf turgor, water balance, photosynthesis performance, and WUE. Thus, it is proposed that beneficial Cl- levels increase the ability of crops to withstand drought, promoting a more sustainable and resilient agriculture.

Chloride Improves Nitrate Utilization and NUE in Plants.

Chloride (Cl-) has traditionally been considered harmful to agriculture because of its toxic effects in saline soils and its antagonistic interaction with nitrate (NO3 -), which impairs NO3 - nutrition. It has been largely believed that Cl- antagonizes NO3 - uptake and accumulation in higher plants, reducing crop yield. However, we have recently uncovered that Cl- has new beneficial macronutrient, functions that improve plant growth, tissue water balance, plant water relations, photosynthetic performance, and water-use efficiency. The increased plant biomass indicates in turn that Cl- may also improve nitrogen use efficiency (NUE). Considering that N availability is a bottleneck for the plant growth, the excessive NO3 - fertilization frequently used in agriculture becomes a major environmental concern worldwide, causing excessive leaf NO3 - accumulation in crops like vegetables and, consequently, a potential risk to human health. New farming practices aimed to enhance plant NUE by reducing NO3 - fertilization should promote a healthier and more sustainable agriculture. Given the strong interaction between Cl- and NO3 - homeostasis in plants, we have verified if indeed Cl- affects NO3 - accumulation and NUE in plants. For the first time to our knowledge, we provide a direct demonstration which shows that Cl-, contrary to impairing of NO3 - nutrition, facilitates NO3 - utilization and improves NUE in plants. This is largely due to Cl- improvement of the N-NO3 - utilization efficiency (NUTE), having little or moderate effect on N-NO3 - uptake efficiency (NUPE) when NO3 - is used as the sole N source. Clear positive correlations between leaf Cl- content vs. NUE/NUTE or plant growth have been established at both intra- and interspecies levels. Optimal NO3 - vs. Cl- ratios become a useful tool to increase crop yield and quality, agricultural sustainability and reducing the negative ecological impact of NO3 - on the environment and on human health.

SILVOLIVE, a Germplasm Collection of Wild Subspecies With High Genetic Variability as a Source of Rootstocks and Resistance Genes for Olive Breeding.

Wild subspecies of Olea europaea constitute a source of genetic variability with huge potential for olive breeding to face global changes in Mediterranean-climate regions. We intend to identify wild olive genotypes with optimal adaptability to different environmental conditions to serve as a source of rootstocks and resistance genes for olive breeding. The SILVOLIVE collection includes 146 wild genotypes representative of the six O. europaea subspecies and early-generations hybrids. These genotypes came either from olive germplasm collections or from direct prospection in Spain, continental Africa and the Macaronesian archipelago. The collection was genotyped with plastid and nuclear markers, confirming the origin of the genotypes and their high genetic variability. Morphological and architectural parameters were quantified in 103 genotypes allowing the identification of three major groups of correlative traits including vigor, branching habits and the belowground-to-aboveground ratio. The occurrence of strong phenotypic variability in these traits within the germplasm collection has been shown. Furthermore, wild olive relatives are of great significance to be used as rootstocks for olive cultivation. Thus, as a proof of concept, different wild genotypes used as rootstocks were shown to regulate vigor parameters of the grafted cultivar "Picual" scion, which could improve the productivity of high-density hedgerow orchards.

Hydraulic Traits Emerge as Relevant Determinants of Growth Patterns in Wild Olive Genotypes Under Water Stress.

The hydraulic traits of plants, or the efficiency of water transport throughout the plant hydraulic system, could help to anticipate the impact of climate change and improve crop productivity. However, the mechanisms explaining the role of hydraulic traits on plant photosynthesis and thus, plant growth and yield, are just beginning to emerge. We conducted an experiment to identify differences in growth patterns at leaf, root and whole plant level among four wild olive genotypes and to determine whether hydraulic traits may help to explain such differences through their effect on photosynthesis. We estimated the relative growth rate (RGR), and its components, leaf gas exchange and hydraulic traits both at the leaf and whole-plant level in the olive genotypes over a full year. Photosynthetic capacity parameters were also measured. We observed different responses to water stress in the RGRs of the genotypes studied being best explained by changes in the net CO2 assimilation rate (NAR). Further, net photosynthesis, closely related to NAR, was mainly determined by hydraulic traits, both at leaf and whole-plant levels. This was mediated through the effects of hydraulic traits on stomatal conductance. We observed a decrease in leaf area: sapwood area and leaf area: root area ratios in water-stressed plants, which was more evident in the olive genotype Olea europaea subsp. guanchica (GUA8), whose RGR was less affected by water deficit than the other olive genotypes. In addition, at the leaf level, GUA8 water-stressed plants presented a better photosynthetic capacity due to a higher mesophyll conductance to CO2 and a higher foliar N. We conclude that hydraulic allometry adjustments of whole plant and leaf physiological response were well coordinated, buffering the water stress experienced by GUA8 plants. In turn, this explained their higher relative growth rates compared to the rest of the genotypes under water-stress conditions.

Silent S-Type Anion Channel Subunit SLAH1 Gates SLAH3 Open for Chloride Root-to-Shoot Translocation.

Higher plants take up nutrients via the roots and load them into xylem vessels for translocation to the shoot. After uptake, anions have to be channeled toward the root xylem vessels. Thereby, xylem parenchyma and pericycle cells control the anion composition of the root-shoot xylem sap [1-6]. The fact that salt-tolerant genotypes possess lower xylem-sap Cl(-) contents compared to salt-sensitive genotypes [7-10] indicates that membrane transport proteins at the sites of xylem loading contribute to plant salinity tolerance via selective chloride exclusion. However, the molecular mechanism of xylem loading that lies behind the balance between NO3(-) and Cl(-) loading remains largely unknown. Here we identify two root anion channels in Arabidopsis, SLAH1 and SLAH3, that control the shoot NO3(-)/Cl(-) ratio. The AtSLAH1 gene is expressed in the root xylem-pole pericycle, where it co-localizes with AtSLAH3. Under high soil salinity, AtSLAH1 expression markedly declined and the chloride content of the xylem sap in AtSLAH1 loss-of-function mutants was half of the wild-type level only. SLAH3 anion channels are not active per se but require extracellular nitrate and phosphorylation by calcium-dependent kinases (CPKs) [11-13]. When co-expressed in Xenopus oocytes, however, the electrically silent SLAH1 subunit gates SLAH3 open even in the absence of nitrate- and calcium-dependent kinases. Apparently, SLAH1/SLAH3 heteromerization facilitates SLAH3-mediated chloride efflux from pericycle cells into the root xylem vessels. Our results indicate that under salt stress, plants adjust the distribution of NO3(-) and Cl(-) between root and shoot via differential expression and assembly of SLAH1/SLAH3 anion channel subunits.