Crops of the future: building a climate-resilient plant immune system

Abstract

A grand challenge facing plant scientists today is to find innovative solutions to increase global crop production in the context of an increasingly warming climate. A major roadblock to global food sufficiency is persistent loss of crops to plant diseases and insect infestations. The United Nations has declared 2020 as the International Year of Plant Health. For historical reasons, molecular studies of plant-biotic interactions in the past several decades have not paid enough attention to how variable climate conditions affect plant-biotic interactions. Here, we highlight a few recent studies that begin to reveal how major climatic drivers impact the plant immune system, particularly secondary messenger and defense hormone signaling, and discuss possible approaches toward engineering climate-resilient plant immunity as part of an ongoing global effort to design “dream” crops of the future.

Introduction

In natural ecosystems and crop fields, plants and their biotic antagonists interact under dynamic environmental conditions. While underlying genetic factors of both the plant host and the pathogen/pest are crucial in shaping the outcome of the interaction, disease outbreak or infestation is often determined by environmental conditions. In the case of a plant-pathogen interaction, a decisive role of environmental conditions in the development of a disease is illustrated in the famous “disease triangle” concept: a disease outbreak requires the interaction of (i) a susceptible host, (ii) a virulent pathogen, and (iii) an environment favorable for disease development [1]. In many cases, adaptation of a plant to various abiotic factors present in the environment can leave it more susceptible to pathogens and pests; however, it is important to note that environmental conditions that negatively impact immunity in one species may not necessarily negatively impact all plant species [2].

Because current climate change can impose suboptimal growth conditions on many crop plants, the environmental impact on plant-biotic interactions is becoming an even more prominent concern in the context of ongoing global climate change [3]. Atmospheric carbon dioxide, temperature, humidity shifts will likely lead to increasing severity of many plant diseases, insect infestations, and migration of important pathogens and insect pests into new geographic ranges [3–5]. Understanding how major climate conditions impact various plant immune pathways has emerged as a timely research topic and is urgently needed to design a new generation of crop plants that can withstand harsh climate conditions under which the current plant immune signaling pathways may fail. Several recent reviews have discussed how various environmental conditions might affect plant-microbe interactions [2,6–10]. In this opinion piece, we aim to highlight several recent studies and discuss possible strategies to counter the negative effects imposed on plant immunity by environmental conditions.

Environmental conditions that modulate plant immunity

A variety of environmental conditions, ranging from light intensity and quality to nutritional status, are known to influence the plant immune response [11,12]. Of these, temperature, CO₂ concentration, and moisture shifts (i.e., drought and high humidity/flooding) are among the most relevant environmental conditions associated with global climate change. We focus our review on how these environmental conditions impact various components of plant immunity (Figure 1).

Figure 1. A simplified diagram depicting the impact of elevated temperature on PTI, ETI, SA and JA immune signaling modules.

Plants can recognize pathogens through (i) cell surface-localized pattern-recognition receptors (PRRs), which perceive conserved microbial patterns, such as bacterial flagellin, and (ii) intracellularly-localized nucleotide-binding, leucine-rich repeat proteins (NLRs), which recognize effector proteins secreted into the plant cell during infection, leading to pattern-triggered immunity (PTI) and effector-triggered immunity (ETI), respectively. Activation of PTI and ETI evokes an increase in secondary messengers, such as reactive oxygen species (ROS) and calcium ions (shown for PTI), and production of SA and subsequent signaling through NPR receptors and TGA transcription factors. As discussed in this paper, elevated temperature (indicated by a thermometer with an upward arrow) has been shown to downregulate ETI, PTI and SA production and signaling (indicated by blue suppression arrows). In contrast, elevated temperature potentiates wound-induced JA signaling through stabilization of the JA receptor coronatine-insensitive 1 (COI1) protein with increased heat shock protein 90 (HSP90) (indicated by a horizontal red arrow). PAMP: Pathogen-associated molecular pattern; ICS1: Isochorismate synthase 1; MAPK: Mitogen-activated protein kinase; SA: Salicylic acid; JA: Jasmonic acid; NPR: Nonexpressor of PR Genes. Created with BioRender.com.

Plants contain two types of immune receptors, which upon activation initiate either pattern-triggered immunity (PTI) or effector-triggered immunity (ETI) [13]. PTI is initiated by cell surface-localized pattern-recognition receptors (PRRs), which recognize conserved microbial patterns, such as bacterial flagellin or fungal chitin [13]. ETI is initiated by mostly intracellularly-localized receptors called nucleotide-binding, leucine-rich repeat proteins (NLRs), which recognize effector proteins that are mostly delivered into the plant cell during pathogen infection [13]. Rapid influx of Ca²⁺ into the cytoplasm is one of the earliest phenomena during both PTI and ETI, occurring after pathogen recognition. The changes in cytosolic Ca²⁺ concentration are thought to be involved in orchestrating downstream immune responses [ 14,15]. Recent studies show that Arabidopsis cyclic nucleotide-gated channel 2/4 (cngc2/cngc4) and hyperosmolality-gated Ca²⁺ permeable channel 1.3/1.7 (osca1.3/osca1.7) mutants exhibit an attenuated cytosolic Ca²⁺ influx in response to the flg22 peptide, a flagellin-derived elicitor of PTI [16•,17•]. The cngc2/cngc4 mutants exhibit reduced immunity against a nonpathogenic mutant (the hrcC mutant defective in bacterial type III secretion) of Pseudomonas syringae pv. tomato (Pst), while the osca1.3/osca1.7 mutant have defects in stomatal immunity against a coronatine-deficient mutant of Pst DC3000 [16•,17•]. Several Ca²⁺-modulated transcription factors, such as calmodulin-binding transcription activators (CAMTAs) and calmodulin-binding protein 60g (CBP60g), are important regulators of PTI and/or ETI [18–21]. Recently, it has been reported that elevated temperature (28 °C) suppresses cytosolic Ca²⁺ influx induced by flg22 in Arabidopsis [22••]. This result suggests that a yet-to-be-identified calcium signal component(s) of the PTI pathway is vulnerable to elevated temperature, even though flg22-induced mitogen-activated protein kinase (MAPK) activity is resilient to elevated temperatures [23]. Previous research has shown that ETI can be suppressed at elevated temperature [6]. In particular, elevated temperature negatively affects stability and/or nuclear localization of several disease resistance proteins [6]. Future research should examine whether ETI-associated Ca²⁺ influx is also affected at elevated temperature and whether PTI and ETI share similar or distinct Ca²⁺ signaling modules.

In addition to Ca²⁺ influx, production of another defense-associated second messenger, reactive oxygen species (ROS), is also affected by environmental conditions. For example, heat shock (30 - 45 min at 42 °C) was recently shown to suppress flg22-induced ROS in Arabidopsis [24•]. Further analysis showed that expression of the FLS2 gene, which encodes the receptor for recognizing the flg22 peptide, was inhibited by heat shock. As a result, the FLS2 protein in the plasma membrane (PM) fraction decreased, indicating that heat shock suppresses the recognition of flg22 by reducing the amount of the FLS2 receptor on the PM [24•].

Beyond Ca²⁺ and ROS, it is well documented that environmental conditions can affect the biosynthesis and/or signaling of several defense-related phytohormones, including salicylic acid (SA) and jasmonic acid (JA). For example, the observation that SA accumulation and signaling induced by pathogens can be negatively affected by elevated temperature has been known for decades. It was reported that the SA production and expression of pathogenesis-related proteins (PRs) induced by tobacco mosaic virus (TMV) that occurs at 20 °C does not occur at 32 °C in the incompatible ‘Samsun NN’ tobacco, which contains the TMV-recognizing N resistance gene [25]. The dramatic impact of elevated temperature on SA production also occurs independently of R gene-mediated resistance (i.e., in the compatible interaction between Arabidopsis and Pst DC3000) [26]. Thus, elevated temperatures appear to have a pervasive negative effect on the production of SA during both incompatible and compatible host-pathogen interactions. As the SA defense pathway is crucial for plant resistance against numerous biotrophic and hemibiotrophic pathogens and insects in crop fields, the down-regulation of SA production at warmer temperatures is a major concern in a warning climate.

In addition to SA, elevated temperature has also been shown to affect the JA pathway during pathogen and insect interactions [26,27•]. Contrary to the effect of elevated temperatures on the SA pathway, JA response gene expression is enhanced at elevated temperatures in Arabidopsis treated with the SA analog benzothiadiazole (BTH), which is used to simulate biotrophic pathogen attacks [26]. Similarly, at elevated temperatures, wound-induced JA responses were potentiated, in this case through increased accumulation of the JA receptor COI1 protein, in both tomato and Arabidopsis. Mechanistically, the accumulation of the COI1 protein is likely mediated by up-regulation of heat shock protein 90 (Hsp90) [27•], as previous V shown for the auxin receptor TIR1 [28]. However, even though wound-induced JA responses are potentiated at elevated temperatures, plant defense against the insect pest Manduka sexta (M. sexta) is still inefficient. This is because elevated temperature potentiates not only JA response, but also insect herbivory, which unfortunately outweighs the enhanced JA response [27•]. Consequently, potentiated wound-induced JA responses were insufficient to counter the more aggressive M. sexta, resulting in exacerbation of the potential detrimental effects of heat waves [27•].

Similar to elevated temperature, elevated CO₂ (eCO₂) impacts plant immunity by modulating normal defense hormone signaling. Specifically, eCO₂ appears to increase basal levels of SA and JA to prime SA- and JA-inducible marker gene expression in Arabidopsis [29]. eCO₂-potentiated immunity against the necrotrophic fungus Plectosphaerella cucumerina (Pc) was not observed in aos1-1 and jar1-1, which have defects in JA production or signaling, respectively, suggesting that eCO₂ affects plant immunity against Pc through JA [29]. Similarly, resistance against brown spot disease in rice, caused by the fungal pathogen Cochliobolus miyabeanus, is enhanced in plants grown under eCO₂ [30]. This elevated resistance is associated with higher activities of rice enzymes involved in ROS scavenging, biosynthesis of phenolic compounds, and increased cell wall reinforcement. However, several reports also indicate that eCO₂ can exhibit profoundly negative impacts on plant defense potentiation [31,32], indicating a need to finely resolve specific environmental and growth conditions that favor defense priming or attenuation in response to excessive CO₂.

High humidity has a profound effect on plant disease development by, for example, favoring the establishment of an aqueous microenvironment for pathogen multiplication [33]. A recent study showed that high humidity can also negatively impact plant immune responses, including bacterium-triggered stomatal closure in both Phaseolus vulgaris and Arabidopsis [34]. In addition, stomatal closure induced by exogenous SA treatment is less efficient and expression of SA responsive genes is down-regulated under high humidity, implying that SA responses in guard cells are dampened under high humidity [34].

Drought is another major issue associated with a warming climate and can exacerbate plant diseases in many cases [35–38]. Drought negatively affects ROS production and expression of defense genes against Magnaporthe oryzae in rice [39]. Individual treatment of plants with either drought or Pst normally causes an upregulation in abscisic acid (ABA), a hormone critical for regulating stomatal aperture [40]. However, under the combined stresses of drought and Pst infection reprogramming of hormonal networks manifests in restrained biosynthesis of ABA [40]. On the other hand, flooding or submergence seem to potentiate some plant immune responses in Arabidopsis and maize [41,42] and, when flooding stress is combined with infestation by the insect pest Spodoptera frugiperda, biosynthesis of SA and related metabolites is increased in maize [42].

Despite the well documented effects of elevated temperature, high humidity and increased CO₂ concentration on SA and JA production and signaling, the exact “immunity targets” of these effects are currently unknown. Identifying these targets and elucidating the underlying mechanisms present an exciting area of future research in the coming decade. In the case of elevated temperature, neither loss of individual hormones that are antagonistic to the SA pathway, such as ABA, JA and ethylene (ET), nor known thermosensory mechanisms, such as those mediated by the PhyB/PIF4 pathway, seems to consistently show a decisive role [26,43], indicating possibly alternative temperature “sensing” and transduction mechanisms in affecting immune responses.

Prospecting wild relatives for engineering climate resilient immunity

As humans selected for desirable agricultural traits (e.g., high seed/biomass yield, seed pod stability, etc.), many characteristics - their natural aegis - that made wild plants resilient to changes in environmental conditions have been lost in domesticated crops [44]. These lost traits are likely tied to what has been termed the dispensable genome, a part of the larger pangenome (the totality of genetic diversity within a species, including its wild relatives) [45,46]. For many years, there has been a vigorous push to dip into this dispensable genome by incorporating more crop wild relatives (CWRs) exhibiting resilience to various abiotic and biotic stresses into breeding programs (https://www.cwrdiversity.org/) [47–50]. The incorporation of CWRs into molecular breeding programs has already led to recent increases in crop yields by up to 30% as new methodologies have accelerated this tedious process [49]. For example, CWRs of both Durham wheat and eggplant have been used for introgression to confer tolerance to multiple climatic-change associated abiotic factors [51,52].

Bolstering the hope of finding climate-resilient defense gene haplotypes, recent de novo resequencing of eight Arabidopsis accessions utilizing the ability to generate and assemble large contigs with PacBio sequencing highlighted that there are many genes (~100-200 genes on average) and structural variants (SVs; e.g., chromosomal inversions, loss or gain of genes) specific to individual accessions [53••]. As these hot spots of genomic rearrangement do not easily undergo meiotic recombination, genomic duplications and rearrangements may drive evolutionary changes in response to selective pressures present in the environment for the rapid development of resistance to biotic stress [46]. Progress is being made to characterize the extent of SVs present in CWRs of some crops (e.g., African and Asian rice and wild soybean) [54–56]. Reconstitution of syntenic regions to allow the introgression of CWR haplotypes via meiosis may not be easy; however, CRISPR/Cas technologies have been developed to force chromosomal inversions [57] or translocations [58•], which affords researchers the ability to create SVs in elite cultivars that could be meiotically compatible with CWRs. Recently, Solanum pimpinellifolium, a CWR of tomato that exhibits tolerance to a wide variety of biotic and abiotic stresses [59], was domesticated de novo via CRISPR-Cas9 technology by editing genes involved in fruit and meristem regulation to those present in domesticated tomato [60]. This exemplifies the potential application of this technology to discover, isolate, and deploy climate resilient traits of CWRs.

CWR-based approaches are likely constrained by germplasm resources. Therefore, the research community now needs to consider innovative methods for developing climate-resilient plants based on the ever increasing knowledge of abiotic and biotic networks in model plants and accessions acquired over the past several decades. One attractive hypothesis is that the complicated climate effects on the plant immune systems may converge onto a few critical transcription factors that regulate biotic and abiotic stresses. If so, identification of these transcription factors could lead to climate-resilient plants via “targeted engineering” using pathogen-dependent uORFs [61•] or CRISPR/Cas-mediated promoter variants [62]. For example, if suppression of SA biosynthesis at elevated temperatures is caused by downregulation of a transcription factor gene, innovative strategies [63•,64] to preserve pathogen-dependent expression of this transcription factor could potentially make SA-mediated immunity resilient to a warm climate.

Targeted genome engineering for climate resilient crops

An important future direction of plant biotechnology is to engineer “dream” crops that exhibit increased biomass or seed yields along with the ability to resist the panoply of biotic and abiotic stresses encountered throughout the plan’s life. While biotechnologies can accelerate the incorporation of loci or traits of biotic and abiotic stress tolerance from pangenome mining expeditions (as discussed above), spatiotemporal regulation of immune programing may be required to compliment gained traits and reduce the impact of growth-defense tradeoffs [65]. Recent studies have employed such a strategy by utilizing an inducible promoter system that finely tunes seed yield and biotrophic defenses. For example, overexpression of Ideal Plant Architecture 1 (IPA1; a member of the plant-specific Squamous-like protein family of transcription factors) in rice causes broad-spectrum resistance to the bacterial pathogen Xanthomonas oryzae pv. oryzae (Xoo) [66]. However, when IPA1 is constitutively expressed at high levels, plant biomass and grain yield decreases, a phenomenon typically observed when defense hormone signaling is constitutively amplified. To obtain an optimal middle ground between growth and defense, rice strains with IPA1 expression under a bacterium-inducible promoter were developed, thusly enabling plants to elevate IPA1 transcription to a very high level only under pathogen attack. This controlled, transient high expression of IPA1 leads to both increased grain yield and bacterial resistance [66]. A similar system utilizing an upstream open reading frame (uORF) system that induces translation of Arabidopsis Nonexpressor of PR Genes 1 (AtNPR1) in rice only under pathogen attack has also been developed to reduce growth-defense tradeoffs [63•]. Screening technologies have been developed to identify biotic or abiotic signals that relieve the suppression of translation imposed by uORFs, which could further facilitate engineering resilience to concomitant biotic and abiotic stresses [61•].

Conclusions and Outlook

In the past four decades, tremendous progress has been made to understand the plant immune system at the molecular level, mostly under plant growth conditions that minimize environmental stresses. However, the dramatic weather conditions associated with the current global climate change highlights a strong need to understand the relationships between the plant immune system and environmental conditions. We argue that, without an aggressive effort to understand such interplays, our knowledge of the plant immune system is incomplete and our ability to develop pathogen/insect/weed-resistant crops in the coming decades will be severely handicapped. Furthermore, while we have learned much from studies performed under well-controlled laboratory conditions, these settings likely limit our understanding of the dynamic plant-biotic interactions that occur in nature and necessitate future studies to be more reflective of the stochastic environments crops encounter. We believe that many climate-resilient points of interplay between the plant immune system and environmental conditions can be found in CWRs, whereas other climate-resilient points need to be engineered based on the current knowledge of the plant immune system in climate-vulnerable plants and accessions. It is an exciting time for plant scientists to make further progress in this field and to push molecular breeding and agricultural biotechnologies toward a new generation of climate-resilient crops for the future (Figure 2).

Figure 2. Building a climate-resilient plant immune system.

Molecular breeding utilizing pangenomic and biotechnological methods, such as CRISPR/Cas9 and uORF-mediated translational control, will allow researchers to develop crops with climate-resilient PTI, ETI, SA and JA signaling modules and other immune responses (not shown). Created with BioRender.com.

Highlights.

• Climate change has a significant impact on the plant immune system.

• Pan-genomes of crop species likely hold important alleles for climate and disease resilience.

• CRISPR-Cas technology may revolutionize crop engineering for climate and disease resilience.