Priming-Mediated Stress and Cross-Stress Tolerance in Crop Plants

By Masayuki Fujita and Bingru Huang

Priming-Mediated Stress and Cross-Stress Tolerance in Crop Plants provides the latest, in-depth understanding of the molecular mechanisms associated with the development of stress and cross-stress tolerance in plants. Plants growing under field conditions are constantly exposed, either sequentially or simultaneously, to many abiotic or biotic stress factors. As a result, many plants have developed unique strategies to respond to ever-changing environmental conditions, enabling them to monitor their surroundings and adjust their metabolic systems to maintain homeostasis. Recently, priming mediated stress and cross-stress tolerance (i.e., greater tolerance to a second, stronger stress after exposure to a different, milder primary stress) have attracted considerable interest within the scientific community as potential means of stress management and for producing stress-resistant crops to aid global food security.

Priming-Mediated Stress and Cross-Stress Tolerance in Crop Plants comprehensively reviews the physiological, biochemical, and molecular basis of cross-tolerance phenomena, allowing researchers to develop strategies to enhance crop productivity under stressful conditions and to utilize natural resources more efficiently. The book is a valuable asset for plant and agricultural scientists in corporate or government environments, as well as educators and advanced students looking to promote future research into plant stress tolerance.

• Provides comprehensive information for developing multiple stress-tolerant crop varieties
• Includes in-depth physiological, biochemical, and molecular information associated with cross-tolerance
• Includes contribution from world-leading cross-tolerance research group
• Presents color images and diagrams for effective communication of key concepts

• Language: English
• Publisher: Academic Press
• Release date: Jan 22, 2020
• ISBN: 9780128178935

Book preview

Agronomy.

Preface

Mohammad Anwar Hossain, Mymensingh, Bangladesh

Fulai Liu, Copenhagen, Denmark

David J. Burritt, Dunedin, New Zealand

Masayuki Fujita, Kagawa, Japan

Bingru Huang, Rutgers, New Brunswick, NJ, United States

Plants growing under field conditions are constantly exposed, either sequentially or simultaneously, to more than one abiotic or biotic stress factor. Plants have developed unique strategies to respond to ever-changing environmental conditions, which enable them to monitor their surroundings and adjust their metabolic systems to maintain homeostasis. Plants acclimate to abiotic and biotic stresses by activating a cascade or network of events that starts with stress perception and ends with the expression of a battery of stress-associated genes. The key components involved in plant stress interactions are the stress stimulus, signal transduction, transcription regulators, target genes, and stress responses, including morphological, biochemical, and physiological changes.

Recently, priming mediated stress and cross-stress tolerance (i.e., greater tolerance to a second stronger stress after exposure to a different more mild primary stress) has attracted considerable interest within the scientific community, as a potential means of stress management and also for producing stress-resistant crops to aid with global food security. Many studies dealing with a range of plant species under different conditions have focused on priming mediated stress and cross-stress tolerance in plants, with recent results indicating that plants have a memory process where a past stress exposure enables them to be better prepared for exposure to stress in the future. While it is known that the induction of cross-tolerance in plants often involves common factors, an in-depth understanding of the molecular mechanisms associated with the development of cross-stress tolerance in plants is still lacking. In addition, to date most of the information obtained on tolerance mechanisms has been obtained from experiments where plants have been exposed to a single form of stress and the mechanisms associated with the tolerance of plants to two or more stresses, to which they are exposed individually or simultaneously, are not fully understood. Hence, there is an urgent need to improve our understanding of the complex mechanisms involved with priming mediated stress cross tolerance, to help develop modern varieties of crop plants that are more resilient to environmental stress.

In this book, we present a collection of 19 chapters written by leading experts researching various aspects of cross-stress tolerance in plants. The aim of this book is to provide a comprehensive overview of the latest advances in our understanding of the physiological, biochemical, and molecular basis of priming mediated stress tolerance and cross-stress tolerance in plants. This will help researchers to develop strategies to enhance crop productivity under stressful conditions and to utilize natural resources more efficiently to ensure future food security. Finally, this book will be a valuable resource for promoting future research into plant stress tolerance, and aims to be a reference book for researchers working on developing plants tolerant to multiple abiotic and biotic stressors.

Chapter 1

Priming mediated stress and cross-stress tolerance in plants: Concepts and opportunities

Eugenio Llorens; Ana I. González-Hernández; Loredana Scalschi; Emma Fernández-Crespo; Gemma Camañes; Begonya Vicedo; Pilar García-Agustín    Group of Biochemistry and Biotechnology, Department of Agricultural Sciences, Universitat Jaume I (UJI) of Castellon, Castellon de la Plana, Spain

Abstract

Plants are often exposed simultaneously to several stresses, and the ability to perceive the stresses and activate the proper responses is crucial for the survival of the plant. Plants possess an intricate network of stress signals including signaling pathways, hormones, and defensive proteins, which activate the most suitable defensive mechanisms against the stresses. This immune system can be stimulated to activate the plant defenses and get them ready for subsequent stresses. In this way, cross-tolerance and priming have emerged as potential solutions for enhancing crop resilience. Cross-tolerance is induced by a mild primary stress, which activates common defenses that reinforce tolerance to different stresses. On the other hand, the application of priming agents prepares the plant to respond faster and more effectively against future stresses. Both mechanisms can improve the resistance of plants against a broad number of stresses simultaneously, providing an effective approach for plant tolerance to environmental perturbations.

Keywords

Cross-tolerance; Priming; Induced resistance; Biotic stress; Abiotic stress

1.1 Introduction: The plant immune system

Plants are not able to escape from the stress factors inherent to the land, for this reason, they have developed a natural immune system, which is usually enough to cope with mild stresses.¹ This immune system includes defensive mechanisms that range from biochemical and molecular signaling to cellular and structural modifications. The capacity of a certain plant to perceive and respond to certain biotic and abiotic stimuli will define its degree of resistance in an ever-changing environment.

1.1.1 Responses against abiotic stress

Abiotic stress is caused by any alteration in the environmental conditions where the plant is growing, and may retard its development. Major abiotic stresses include high and low temperatures, drought, flood, light, radiation, salinity, heavy metals, or alterations in the level of nutrients. This broad spectrum of stressors entails that more than 95% of land area worldwide is affected by a certain abiotic stress.² The response to these environmental perturbations is related to specific changes in the gene expression and the physiology of the plant. After the perception of the stress, plants generate reactive oxygen species (ROS) that can act as early response signaling molecules. Elevated ROS levels can induce the accumulation of phytohormones such as abscisic acid (ABA) or gibberellins (GA). Specifically, ABA has a main role in the response against abiotic stress such as water deficit and salinity, where it regulates the aperture of stomata and induces the expression of ABA-responsive genes.³,⁴ However, the response against drought and salinity, as well as cold stress, could also be mediated by an ABA-independent pathway.⁵

1.1.2 Responses against biotic stress

Plants possess two types of defenses against pests and pathogen attacks. On one hand, the constitutive defenses are preformed barriers that bring the first layer of defense. This layer includes physical defenses such as wax and thickened cuticles, but also a constitutive accumulation of chemical deterrents such as pyrethrins, phytoalexins, and phytoanticipins.⁶ On the other hand, plants are able to recognize the presence of pests and microorganisms and trigger a defensive response accordingly. The detection of the pathogen starts with the perception of the microbial- or pathogen-associated molecular patterns (MAMPs or PAMPs), which are small molecules or molecular motives conserved within a class of microbes.¹ The PAMPs are recognized by receptors localized at the surface of the cells called pattern recognition receptors (PRR). If the plant is able to recognize the PAMP successfully, it will trigger the first layer of induced defense called PAMP-triggered immunity (PTI). However, pathogens are able to release effector proteins that interfere with PTI, resulting in the so-called effector-triggered susceptibility (ETS).⁷ Once they have entered the tissue, the effectors manipulate the host cell to benefit the infection. At this stage, a battery of intracellular nucleotide-binding/leucine-rich-repeat (NLR) receptors will detect the given effector resulting in the activation of effector-triggered immunity (ETI).¹,⁸ Usually, the ETI is a stronger PTI response coupled with a hypersensitive response and localized cell death that effectively protects the plant from the threat. Both PTI and ETI activate signaling pathways mediated by the accumulation of phytohormones such as salicylic acid (SA), jasmonic acid (JA), and ethylene (ET), which will trigger the ulterior induced defensive responses. Depending on pathogen lifestyle, SA is often related to the resistance against biotrophic and hemibiotrophic pathogens, while JA and ET are related with defense against necrotrophic pathogens and herbivores.⁹

1.2 Induced resistance: Concepts and terminology

Vallad and Goodman¹⁰ described the induced resistance as a physiological state of enhanced defensive capacity elicited by specific environmental stimuli, whereby the plant's innate defenses are potentiated against subsequent biotic challenges. Thereafter, it was demonstrated that the induced resistance can be also effective against abiotic stress. In this way, after the recognition of the stress, besides the local response, the plant is able to induce defensive responses in systemic tissues, protecting in this way parts yet unexposed to the damage. Moreover, it has been demonstrated that the defensive signaling can travel to other plants by the emission of volatile compounds (VOCs), preparing the neighbor plants for an upcoming attack.¹¹ In recent years, it has been demonstrated that this enhanced physiological state can be induced also by mild stress, by the application of natural and chemical compounds, as well as by beneficial microbes.¹²,¹³

1.2.1 Systemic acquired resistance, induced systemic resistance, and systemic acquired acclimation

Depending on the stimuli that induce the systemic response in the plant, the induced resistance has been described as systemic acquired resistance (SAR), induced systemic resistance (ISR), and systemic acquired acclimation (SAA).¹⁴,¹⁵ Whereas SAR and ISR are induced against biotic stresses, SAA is activated in response to abiotic stimuli. SAR is a class of induced resistance characterized by its broad-spectrum against pathogens and durability over time and generations. This systemic resistance can be activated in the plant by the attack of biotrophic pathogens or by treatment with chemical or natural compounds such as benzo(1,2,3)-thiadiazole-7-carbothioic acid S-methyl-ester (benzothiadiazole (BTH); acibenzolar-S-methyl (ASM)) or methyl salicylate.¹⁶ Usually, SAR is defined as the induced resistance mediated by the accumulation of SA, which includes the subsequent expression of PR genes; whereas ISR is often associated with the defensive responses mediated by JA and ET. Initially, ISR was described as a resistance induced by beneficial microbes such as rhizobacteria. However, in later studies, it was observed that it can be also induced by treatments with chemical and natural compounds and by the attack of pathogens.¹⁷

SAA is described as the systemic acclimation induced by an abiotic stress in parts of the plant that are not yet exposed.¹⁴ Moreover, recent studies suggested that SAA is able to induce resistance also against biotic stresses.¹⁸ Although the mechanism of SAA is not fully described, several studies have suggested that SAA is activated against heat, salinity, and high light stress, including the activation of an ROS wave.¹⁹,²⁰ However, other factors mediating specificity to SAA response have been described, such as calcium waves, electric signals, and hormones.¹⁴,²¹

1.2.2 Induction of resistance: Cross-tolerance and priming

In the field, plants are continuously exposed to a broad range of stresses that can occur either simultaneously or successively. Plants respond to stress by activating a variety of responses at a molecular, cellular, and physiological level that lead to tolerance or resistance mechanisms. Recently, accumulated evidence has shown that plants exposed to a mild primary stress can acquire resistance to a second, strong stress. This phenomenon is known as cross-tolerance, cross-resistance, or multiple-stress resistance and allows plants to acquire tolerance to a broad range of stresses.²²

The exposure to a certain stress can induce a stress memory, which is persistent and prepares the plant for better and faster response in later stress events in a phenomenon known as priming, which covers three different phases, pre-challenge priming state, challenge priming state, and transgenerational priming state.²³–²⁶ In this case, the priming can be active against both biotic and abiotic stresses. Moreover, some studies suggest that priming can involve epigenetic changes that could be heritable.²⁷ The main characteristic of priming is the lag phase, which separates the priming activation event from the second stress.²⁸ The priming stimuli induce changes in the plant at a biochemical, molecular, and epigenetic level that will be crucial for the defensive responses against the second stress. However, in contrast to a complete defensive response, the changes maintained in the lag phase have low fitness costs. In this way, any stress that induces resistance to a second different stress after a lag phase would be defined as priming induced cross-tolerance.²²

Interestingly, priming of defense can also be induced by natural or chemical elicitors in absence of stress. Compounds such as hexanoic acid (Hx), BTH, or probendazole are able to induce the priming effects, generating changes that remain in the lag phase to boost the defensive responses without causing stress to the plant.²³ However, in this case, since the stimulus is not caused by a stress, this mechanism could not be considered as cross-tolerance, but priming induced resistance.

1.3 Cross-tolerance

In this section, the cross-tolerance mechanism is discussed in detail. Cross-tolerance can be divided into inherent cross-tolerance and induced cross-tolerance. Inherent cross-tolerance can be defined as the capacity of an individual within a species to resist multiple stresses due to its genetic background. This enhanced capacity could be the consequence of a variety of mechanisms involving changes in plant and cell architecture, signal transduction, or detoxification process, as previously mentioned by Poland et al.²⁹ The second type of cross-tolerance, induced cross-tolerance, can be defined as the activation of systemic plant tolerance through previous exposure to another type of stress. Related to this, it has been observed that the tolerance to the subsequent stress is affected by the intensity or duration of the primary stress. In this way, when the intensity of the primary stress exceeds a certain threshold, the organism requires a period of recovery between the first and the second stress to develop effective tolerance.³⁰ For this reason, although research in model organisms suggests that cross-tolerance could be useful for agronomy and breeding of crop plants, only a few researches has been carried out under field conditions.³¹ Therefore, several important issues such as the timing, duration, and intensity of a stressor, as well as its interactions with other biotic and abiotic factors still need to be addressed.

Cross-tolerance has been shown for different types of stress.³² However, among them, cold or heat-shock induced cross-tolerance to abiotic stresses is the most common as highlighted by Hossain et al.²² Yasuda³³ reviewed the advances made to improve cold tolerance in rice at different developmental stages after exposure to heat-shock. Moreover, Sekara et al.³⁴ showed that chilling or heat stress applied to eggplant (Solanum melongena) after germination, at the radical stage, could result in better plant acclimation to chilling at a later stage. Stress cross-tolerance was also reported in Camelia sinensis, since plants subjected to cold stress showed delayed drought-induced leaf senescence mainly due to their enhanced antioxidant capacity.³⁵ A similar stress cross-tolerance was recently reported in Coffea spp., where water shortage in the cold season helped to mitigate cold impact due to an increase in plant antioxidative defenses.³⁶ In relation to abiotic stresses, it was also demonstrated that wounding increased salt stress tolerance in tomato,³⁷ whereas UV radiation enhanced drought tolerance in Pisum sativum and Triticum aestivum.³⁸ In addition, heat shock improved tolerance of maize to heat, chilling, salt and drought³⁹ was also proved.

syndrome, including several symptoms such as leaf chlorosis, lower cation content, or several metabolite changes., as the sole N source, is able to alleviate the effects of high salinity in several plants such as barley, halophytes, or citrus plants¹⁸,⁴¹,⁴² and produce resistance against Pseudomonas syringae in tomato plants,⁴³ inducing several changes in hormones, polyamines, and the antioxidant enzyme machinery. Moreover, Foyer et al.⁴⁴ reviewed how the abiotic environment influences plant responses to attack by phloem-feeding aphids. Related to this, Gonzalez et al.⁴⁵ showed, through metatranscriptomics analysis, that willow trees cultivated on petroleum hydrocarbon contaminated soil display less Tetranychus urticae infestation, while Arasimowicz-Jelonek et al.⁴⁶ revealed that the enhanced defense responses observed in susceptible potato cutivar (Solanum tuberosum) undergoing aluminum (Al) stress at the root level correlated with reduced disease symptoms after leaf inoculation with Phytophthora infestans. In addition, Cu-heptagluconate treatment induces resistance against P. syringae in tomato plants accompanied by a reduction in the amount of ROS and an accumulation of caffeic and chlorogenic acids in infected plants.⁴⁷ Furthermore, drought stress resulted in a reduction in Botrytis cinerea infection as well as a suppression of Oidium neolycopersici in tomato,⁴⁸ whereas, salt-induced osmotic stress was correlated with resistance to powdery mildew in barley.⁴⁹ Besides, exposure to ozone was also shown to induce resistance in Arabidopsis and in tobacco plants against P. syringae strains and tobacco mosaic virus, respectively.⁵⁰,⁵¹

Interestingly, some fungicides and herbicides also appear to act as cross-tolerance inducers. dos Santos et al.⁵² demonstrated that independent or simultaneous application of glyphosate and carfentrazone-ethyl herbicide drift reduced infection and uredinial formation of Austropuccinia psidii on Eucalyptus grandis. Application of the diphenyl ether herbicide Lactofen was also found to reduce disease severity in soybean plants infected by Sclerotinia sclerotiorum. This reduction was accompanied by an increased expression of PR and thaumatin/osmotin-like proteins.⁵³ Moreover, treatment of tobacco with the fungicide pyraclostrobin enhanced resistance to both tobacco mosaic virus and Pseudomonas spp. infection.⁵⁴ Fungicide application not only induces resistance against biotic stress but also against abiotic stress, as demonstrated by Hassan,⁵⁵ who showed that dual application of the fungicide chlorothalonil and the antiozonant compound ethylenediurea (EDU) resulted in higher resistance to O3, probably due to elevated levels of glutathione.

On the other hand, biotic stress has also been reported to increase resistance to abiotic stress. For example, infection of Arabidopsis with the soil borne fungal pathogen Verticillium longisporum (thale cress wilt) resulted in de novo xylem formation, enhancing drought tolerance.⁵⁶ Similarly, Xu et al.⁵⁷ showed that viral infection can induce drought tolerance in several plants species, which correlated with an increase in several osmoprotectants and antioxidants.

Biotic factors can also have a major influence on plant response to other biotic stresses. For instance, herbivore-induced wounding was shown to induce resistance against the necrotrophic B. cinerea through herbivory-induced priming of JA responses.⁵⁸ However, these effects are difficult to predict since they depend on a bunch of factors such as severity and duration of the infection, the lifestyle of the pathogen, the plant species that is attacked, and the biology of the insect. Since SA accumulation was observed in plants infected by biotrophic or hemibiotrophic pathogens, SA treatment, which mimics this accumulation, were performed in tomato plants, increasing their resistance to the hemibiotroph Fusarium oxysporum.⁵⁹

Although cross-tolerance is a phenomenon that often occurs in plants, little is known about its molecular mechanisms. However, it is now clear from studies on single stresses that plant responses to both biotic and abiotic stresses are mediated by common signals such as reactive oxygen and nitrogen species (ROS and RNS, respectively),⁶⁰ calcium gradients,⁶¹ heat shock proteins,⁶² and plant hormones (mainly SA, JA, and ABA).⁶³ The interplay between these signals is considered a key component controlling cross-tolerance.⁶⁴ These mechanisms will be discussed in more detail in the further chapters of this book.

1.4 Priming

As mentioned above, the main characteristic of priming that differentiates it from cross tolerance is the lag phase that separates the priming activation event from the second stress. Besides, the changes that occur in primed plants do not imply a significant fitness cost. Priming can be applied to plants at different growth stages and also in the seeds,⁶⁵ which confers resistance against abiotic and biotic stresses and even a transgenerational resistance.⁶⁶,⁶⁷

Plants can be primed by both abiotic and biotic stimuli. Abiotic stresses can cause similar effects at biochemical, cellular, and molecular levels that are interconnected and activating similar signaling cascades. Therefore, a mild or a short pretreatment with an abiotic stress can enhance tolerance to a second stress. If the stimulus and the stress are the same it is defined as cis-priming, whereas when the stimulus is different from the stress it is known as trans-priming.⁶⁸

The main abiotic stimuli for which priming has been studied are high temperatures, cold, drought, salinity, and chemical compounds. Priming of the plant by high temperatures against a subsequent emergence of the same stress has been widely investigated and represents a clear example of cis-priming. Related to this, it has been demonstrated that heat priming could effectively improve thermo-tolerance to the later recurred heat stress in several plant species by improving the osmoregulation, detoxification, and protection of proteins.⁶⁹,⁷⁰ Moreover, since flowering and fruit development are the phenological stages during which the crop yield is determined, studies have shown that heat priming applied to wheat plants at pre-anthesis stage alleviated the negative effects of postanthesis heat stress on grain yield and quality.⁷¹ The enhanced tolerance to postanthesis heat stress is attributed to priming induced enhanced carbohydrate remobilization from stems to grains resulting in less changed starch content. A priming effect was also observed when high temperatures were applied during the vegetative stage of wheat plants.⁷¹ In this case, primed plants showed higher photosynthesis rate and a better redox homeostasis in relation to the nonprimed plants.

It has also been demonstrated that heat priming in the parental generation can induce transgenerational thermo-tolerance. Related to this, Wang et al.⁷² showed that this might be an effective measure to cope with severe heat stresses during critical growth stages in wheat production. The transgenerational stress tolerance to heat stress in the successive generation was accompanied by higher grain yield, better maintenance of leaf photosynthesis, and enhanced activities of antioxidant enzymes and reduced cell membrane damage. On the other hand, it has been shown that high temperatures could also induce priming in fungi⁷³ or even in bacteria.⁷⁴ These findings support the idea that pathogens could also adapt to an increase in temperature, which is an important factor for understanding the evolution of the host-pathogen system when facing this type of stress.

Cold is another factor that affects plant development and crop production. Its effects, both in terms of sensitivity and acclimation to cold, have been described in detail in the recent review of Baier et al.⁷⁵ Similar to what occurs when plants are subjected to a short stress by high temperatures, short exposure to cold can prime plants against future cold stress events. Cold priming implies accumulation of osmolytes, such as sucrose and proline, which contribute to the stabilization of the membrane.⁷⁶ In Arabidopsis, pretreatment with cold promotes priming effects that maintains photosynthetic performance and reproductive development under a second cold stress.⁷⁷

Besides heat and cold, drought was also shown to induce priming. The review of Wang et al.⁷⁸ describes how several authors have managed to increase the tolerance to water limiting conditions through drought priming. This improved performance in plants was accompanied by an enhancement in photosynthetic and ROS scavenging capacity. Moreover Ben Abdallah et al.⁷⁹ proposed the application of drought stress for priming of Olea europea young plants in order to help them overcome a later drought stress. After the second drought stress, higher integrity of photosystem II, higher proline and sugar accumulation, and a more activated antioxidant system were observed in primed plants as compared with the nonprimed plants. In this way, Wang et al.⁷¹ showed that in wheat, drought priming before anthesis improved the tolerance to a subsequent drought stress during grain filling, resulted in higher grain yield and enhanced photosynthesis rate and ascorbate peroxidase activity in the primed plants than in the nonprimed plants. In addition, it was also demonstrated that drought priming applied to wheat plants during the terminal growth stage together with seed osmotic and hydropriming improved plants transgenerational salt tolerance⁸⁰ and transgenerational drought tolerance⁸¹ by modulating the water relations, osmolytes accumulation, malondialdehyde contents, and lipid peroxidation.

Halopriming examples can also be found; however, these treatments were mostly performed in seeds rather than in plants. Related to halopriming in plants, Caparrotta et al.⁸² showed that Vicia faba plants located in the proximity of salt primed plants were more tolerant to a later salt stress due to VOCs signals, as exemplified by the analysis of plant growth, osmotic adjustment, leaf gas exchanges, and chlorophyll fluorescence. Moreover, it has been demonstrated in other works that salt priming combined with biocontrol agents can induce a priming state required for triggering an early expression of plant defense genes against Pyricularia oryzae.⁸³ Seed halopriming has been carried out in several crops such as wheat,⁸⁴ inducing antioxidative responses against salinity; in melon,⁸⁵ inducing osmoregulation by the accumulation of organic solutes; in sunflower,⁸⁶ inducing salt tolerance by osmoregulation; and in sugarcane,⁸⁷ where the plants that developed from the primed seeds showed a major tolerance to future salinity and drought stresses.

Savvides et al.,⁸⁸ in their recent review, described how certain chemical compounds primed plants against multiple abiotic stresses. Among these compounds, hormones, proline, melatonin, polyamines, and reactive oxygen-nitrogen-sulfur species (RONSS: NO donors, H2O2, H2S donors) induced activation of signaling pathways that potentially resulted in the systemic accumulation of dormant tolerance signals in the primed plants. When facing a future stress, these plants showed enhanced stress tolerance-related responses such as improved ROS detoxification, osmoprotection, protein stabilization, and ion homeostasis.

Trans-priming by chemical compounds against heat was documented in different works. Methyl jasmonate treatment of Arabidopsis before heat stress conferred protection to the plants and revealed the role of oxylipin pathway against this stress.⁸⁹ Moreover, melatonin pretreatment of tomato plants showed a priming effect against heat stress by promoting cellular protein protection.⁹⁰ Recently, chemical priming of the seeds has also been carried out. For instance, tomato seeds primed with β-sitosterol or gibberellic acid, showed enhanced tolerance against both high and low temperatures.⁹¹

Plants memorize the previous activation of plant immunity when they are exposed to elicitors—this has also been referred to as plant stress memory and defense priming⁹²—and become primed to activate more rapid and/or stronger defense responses following attack by pathogens or insects.²⁵,²⁸,⁹³ Resistance elicitors, also known as priming agents, can be natural (see review by Aranega-Bou et al.⁹⁴), such as hexanoic acid, which induces plant defenses by means of priming mechanism that acts differentially, depending on the pathogen,⁹⁵–⁹⁷ or synthetic compounds, such as 2,6-dichloroisonicotinic acid⁹⁸ or BTH.⁹⁹ Moreover, virulent/avirulent pathogens¹⁰⁰ or beneficial microbes,¹⁰¹ nonpathogenic rhizobacteria, and mycorrhizal fungi¹⁰²,¹⁰³ can provoke the same priming effect. Likewise, beneficial microbes must suppress local immune responses in the host to establish the symbiosis through the secretion of compounds with eliciting activity. There are many studies providing evidence that the induced resistance by this method is based on priming mechanism (see review published by Pieterse et al.¹⁷). Besides, arthropod attack can also produce priming in plants due to oral secretions when plants are bitten by the insect,¹⁰⁴ oviposition,¹⁰⁵ or physical contact, since trichomes can perceive insect contact and prepare the plant to defend subsequent attack¹⁰⁶ and/or produce VOCs. VOCs are emitted by herbivore-infested plants and some of them can prime JA-dependent defenses in systemic tissues and neighboring plants.¹⁰⁷

The mechanisms that are activated by applying a priming agent are according to the challenger life-style. These mechanisms are described in detail in the recent review by Mauch-Mani et al.⁶⁷

In general, a few seconds or minutes after the stimulus induced by the priming agent, transient changes are observed in the level of intracellular calcium. This can be observed in priming by wounding¹⁰⁸ and during arbuscular mycorrhiza fungi (AMF) root colonization.¹⁰⁹ The increase in cytosolic calcium may trigger a membrane depolarization and the ROS burst.¹⁰⁸ A fine-tuning of ROS homeostasis seems to be essential for priming. In a review paper, González-Bosch¹¹⁰ highlighted how the priming agents modulate the oxidative environment and interacting with hormone signaling pathways such as SA, JA, and ET. Moreover, it is widely accepted that a transcriptional and metabolic reprogramming may produce a differential biosynthesis of secondary metabolites as a characteristic process involved during initial priming stage.¹¹¹

Beneficial microorganisms (endophytes and mycorrhizae) induce ISR associated with priming of JA-dependent defenses.¹¹²–¹¹⁴ It is known that the colonization of several endophytes, which involves repression of SA signaling and promotion of JA responses, activates priming against insect attack. The review by Bastias et al.¹¹⁵ showed that Epichloë fungal endophytes improve antiherbivore defenses of their grass hosts via alkaloid-dependent and -independent mechanisms, as well as by inducing JA pathways. Moreover, some authors have shown the ability of endophytic bacteria to induce priming by activating the SA and JA/ET signaling pathways, and by producing resistance-conferring VOCs or disrupting quorum sensing establishment in the