WRKY48 negatively regulates plant acclimation to a combination of high light and heat stress.

Plants growing under natural conditions experience high light (HL) intensities that are often accompanied by elevated temperatures. These conditions could affect photosynthesis, reduce yield, and negatively impact agricultural productivity. The combination of different abiotic challenges creates a new type of stress for plants by generating complex environmental conditions that often exceed the impact of their individual parts. Transcription factors (TFs) play a key role in integrating the different molecular signals generated by multiple stress conditions, orchestrating the acclimation response of plants to stress. In this study, we show that the TF WRKY48 negatively controls the acclimation of Arabidopsis thaliana plants to a combination of HL and heat stress (HL + HS), and its expression is attenuated by jasmonic acid under HL + HS conditions. Using comparative physiological and transcriptomic analyses between wild-type and wrky48 mutants, we further demonstrate that under control conditions, WRKY48 represses the expression of a set of transcripts that are specifically required for the acclimation of plants to HL + HS, hence its suppression during the HL + HS stress combination contributes to plant survival under these conditions. Accordingly, mutants that lack WRKY48 are more resistant to HL + HS, and transgenic plants that overexpress WRKY48 are more sensitive to it. Taken together, our findings reveal that WRKY48 is a negative regulator of the transcriptomic response of Arabidopsis to HL + HS and provide new insights into the complex regulatory networks of plant acclimation to stress combination.

Omics analyses in citrus reveal a possible role of RNA translation pathways and Unfolded Protein Response regulators in the tolerance to combined drought, high irradiance, and heat stress.

Environmental changes derived from global warming and human activities increase the intensity and frequency of stressful conditions for plants. Multiple abiotic factors acting simultaneously enhance stress pressure and drastically reduce plant growth, yield, and survival. Stress combination causes a specific stress situation that induces a particular plant response different to the sum of responses to the individual stresses. Here, by comparing transcriptomic and proteomic profiles to different abiotic stress combinations in two citrus genotypes, Carrizo citrange (Citrus sinensis × Poncirus trifoliata) and Cleopatra mandarin (Citrus reshni), with contrasting tolerance to different abiotic stresses, we revealed key responses to the triple combination of heat stress, high irradiance and drought. The specific transcriptomic response to this stress combination in Carrizo was directed to regulate RNA metabolic pathways and translation processes, potentially conferring an advantage with respect to Cleopatra. In addition, we found endoplasmic reticulum stress response as common to all individual and combined stress conditions in both genotypes and identified the accumulation of specific groups of heat shock proteins (HSPs), such as small HSPs and HSP70s, and regulators of the unfolded protein response, BiP2 and PDIL2-2, as possible factors involved in citrus tolerance to triple stress combination. Taken together, our findings provide new insights into the acclimation process of citrus plants to multiple stress combination, necessary for increasing crop tolerance to the changing climatic conditions.

Reduction of heat stress pressure and activation of photosystem II repairing system are crucial for citrus tolerance to multiple abiotic stress combination.

Drought, heat and high irradiance are abiotic stresses that negatively affect plant development and reduce crop productivity. The confluence of these three factors is common in nature, causing extreme situations for plants that compromise their viability. Drought and heat stresses increase the saturation of the photosystem reaction centers, increasing sensitivity to high irradiance. In addition, these stress conditions affect photosystem II (PSII) integrity, alter redox balance of the electron transport chain and decrease the photosynthetic rate. Here, we studied the effect of the stress combinations on the photosynthetic apparatus of two citrus genotypes, Carrizo citrange (Citrus sinensis × Poncirus trifoliata) and Cleopatra mandarin (Citrus reshni). Results obtained showed that physiological responses, such as modulation of stomatal aperture and transpiration rate, aimed to reduce leaf temperature, are key to diminishing heat impact on photosynthetic apparatus and increasing tolerance to double and triple combinations of drought, high irradiance and high temperatures. By using transcriptomic and proteomic analyses, we have demonstrated that under these abiotic stress combinations, Carrizo plants were able to increase expression of genes and proteins related to the photosystem repairing machinery (which better maintained the integrity of PSII) and other components of the photosynthetic apparatus. Our findings reveal crucial physiological and genetic responses in citrus to increase tolerance to the combination of multiple abiotic stresses that could be the basis for breeding programs that ensure a sustainable citrus production.

Plant responses to climate change: metabolic changes under combined abiotic stresses.

Climate change is predicted to increase the frequency and intensity of abiotic stress combinations that negatively impact plants and pose a serious threat to crop yield and food supply. Plants respond to episodes of stress combination by activating specific physiological and molecular responses, as well as by adjusting different metabolic pathways, to mitigate the negative effects of the stress combination on plant growth, development, and reproduction. Plants synthesize a wide range of metabolites that regulate many aspects of plant growth and development, as well as plant responses to stress. Although metabolic responses to individual abiotic stresses have been studied extensively in different plant species, recent efforts have been directed at understanding metabolic responses that occur when different abiotic factors are combined. In this review we examine recent studies of metabolomic changes under stress combination in different plants and suggest new avenues for the development of stress combination-resilient crops based on metabolites as breeding targets.

γ-Aminobutyric acid plays a key role in plant acclimation to a combination of high light and heat stress.

Plants are frequently subjected to different combinations of abiotic stresses, such as high light (HL) intensity, and elevated temperatures. These environmental conditions pose a threat to agriculture production, affecting photosynthesis, and decreasing yield. Metabolic responses of plants, such as alterations in carbohydrates and amino acid fluxes, play a key role in the successful acclimation of plants to different abiotic stresses, directing resources toward stress responses, and suppressing growth. Here we show that the primary metabolic response of Arabidopsis (Arabidopsis thaliana) plants to HL or heat stress (HS) is different from that of plants subjected to a combination of HL and HS (HL+HS). We further demonstrate that the combined stress results in a unique metabolic response that includes increased accumulation of sugars and amino acids coupled with decreased levels of metabolites participating in the tricarboxylic acid cycle. Among the amino acids exclusively accumulated during HL+HS, we identified the nonproteinogenic amino acid γ-aminobutyric acid (GABA). Analysis of different mutants deficient in GABA biosynthesis (GLUTAMATE DESCARBOXYLASE 3 [gad3]) as well as mutants impaired in autophagy (autophagy-related proteins 5 and 9 [atg5 and atg9]), revealed that GABA plays a key role in the acclimation of plants to HL+HS, potentially by promoting autophagy. Taken together, our findings identify a role for GABA in regulating plant responses to combined stress.

Citrus rootstocks modify scion antioxidant system under drought and heat stress combination.

KEY MESSAGE: The activation of the antioxidant system under stress combination is a transmissible trait from the rootstock to the scion. Therefore, rootstock selection is key to improve crop performance and a sustainable production under changing climate conditions. Climate change is altering weather conditions such as mean temperatures and precipitation patterns. Rising temperatures, especially in certain regions, accelerates soil water depletion and increases drought risk, which affects agriculture yield. Previously, our research demonstrated that the citrus rootstock Carrizo citrange (Citrus sinensis × Poncirus trifoliata) is more tolerant than Cleopatra mandarin (C. reshni) to drought and heat stress combination, in part, due to a higher activation of the antioxidant system that alleviated damage produced by oxidative stress. Here, by using reciprocal grafts of both genotypes, we studied the importance of the rootstock on scion performance and antioxidant response under this stress combination. Carrizo rootstock, under stress combination, positively influenced Cleopatra scion by reducing H2O2 accumulation, increasing superoxide dismutase (SOD) and ascorbate peroxidase (APX) enzymatic activities and inducing SOD1, APX2 and catalase (CAT) protein accumulations. On the contrary, Cleopatra rootstock induced decreases in APX2 expression, CAT activity and SOD1, APX2 and CAT contents on Carrizo scion. Taken together, our findings indicate that the activation of the antioxidant system under stress combination is a transmissible trait from the rootstock to the scion and highlight the importance of the rootstock selection to improve crop performance and maintain citrus yield under the current scenario of climate change.

High temperatures modify plant responses to abiotic stress conditions.

Climate change is altering environments in which plants and different crops grow and survive. We already experienced an increase in worldwide average earth surface temperatures, as well as frequency and extent of damaging heat waves. These conditions collide in the field with other abiotic stresses such as water deficit, high salinity, increased light irradiation, and so on, generating complex harmful conditions that destabilize agricultural systems. The conditions generated during these episodes of stress combination greatly differ from those occurring in the field when different stress factors occur individually; conditions that have been the focus of study for decades. Fortunately, knowledge of physiological and molecular responses to stress combinations and the cost they inflict on plant growth and yield has been exponentially increasing in the past several years. Understanding plant performance under multiple stress combinations will allow breeding crops capable of maintaining yield production under the new climatic conditions. Here, after reviewing recent data on physiological, hormonal and transcriptional responses to different stress combinations, we highlight the importance of photodamage avoidance, abscisic and jasmonic acid signaling, and the upregulation of genes involved in oxidation-reduction processes, photosynthesis and protein metabolism, for plant acclimation to conditions of high temperatures, in combination with other common abiotic stress factors such as drought or salinity. Finally, we propose new approaches to investigate the response of plants to stress combinations and discuss strategies for improving crop resilience to stress combination.

Jasmonic Acid Is Required for Plant Acclimation to a Combination of High Light and Heat Stress.

In the field, plants experience high light (HL) intensities that are often accompanied by elevated temperatures. Such conditions are a serious threat to agriculture production, because photosynthesis is highly sensitive to both HL intensities and high-temperature stress. One of the potential cellular targets of HL and heat stress (HS) combination is PSII because its degree of photoinhibition depends on the balance between the rate of PSII damage (induced by light stress), and the rate of PSII repair (impaired under HS). Here, we studied the responses of Arabidopsis (Arabidopsis thaliana) plants to a combination of HL and HS (HL+HS) conditions. Combined HL+HS was accompanied by irreversible damage to PSII, decreased D1 (PsbA) protein levels, and an enhanced transcriptional response indicative of PSII repair activation. We further identified several unique aspects of this stress combination that included enhanced accumulation of jasmonic acid (JA) and JA-Ile, elevated expression of over 2,200 different transcripts that are unique to the stress combination (including many that are JA-associated), and distinctive structural changes to chloroplasts. A mutant deficient in JA biosynthesis (allene oxide synthase) displayed enhanced sensitivity to combined HL+HS and further analysis revealed that JA is required for regulating several transcriptional responses unique to the stress combination. Our study reveals that JA plays an important role in the acclimation of plants to a combination of HL+HS.

High temperatures change the perspective: Integrating hormonal responses in citrus plants under co-occurring abiotic stress conditions.

Plants growing in the field are subjected to multiple stress factors acting simultaneously. Abnormally high temperatures are expected to affect wild plants and crops in the next years due to global warming. In this work, we have studied physiological, hormonal and molecular responses of the citrus rootstock, Carrizo citrange (Poncirus trifoliata L. Raf. × Citrus sinensis L. Osb.) subjected to wounding or high salinity occurring individually or in combination with heat stress. According to our results, combination of high salinity and heat stress aggravated the negative effects of salt intoxication in Carrizo. The high transpiration rate caused by high temperatures counteracted physiological responses of plants to salt stress and increased Cl- intake in leaves. In addition, 12-oxo-phytodienoic acid accumulated specifically under combination of wounding and heat stress, whereas at low temperatures, wounded plants accumulated jasmonic acid (JA) and JA-isoleucine (JA-Ile). Moreover, an antagonism between salicylic acid (SA) and JA was observed, and wounded plants subjected to high temperatures did not accumulate JA nor JA-Ile whereas SA levels increased (via isochorismate synthase biosynthetic pathway). Wounded plants did not accumulate abscisic acid (ABA) but its catabolite phaseic acid. This could act as a signal for the upregulation of (ABA)-RESPONSIVE ELEMENT (ABRE)-BINDING TRANSCRIPTION FACTOR 2 (CsAREB2) and RESPONSIVE TO DISSECATION 22 (CsRD22) in an ABA-independent way. This work uncovers some mechanisms that explain Carrizo citrange tolerance to high temperatures together with different hormonal signals in response to specific stresses. It is suggested that co-occurring abiotic stress conditions can modify (either enhance or reduce) the hormonal response to modulate specific responses.

Involvement of ascorbate peroxidase and heat shock proteins on citrus tolerance to combined conditions of drought and high temperatures.

Usually several environmental stresses occur in nature simultaneously causing a unique plant response. However, most of the studies until now have focused in individually-applied abiotic stress conditions. Carrizo citrange (Poncirus trifoliata L. Raf. X Citrus sinensis L. Osb.) and Cleopatra mandarin (Citrus reshni Hort. ex Tan.) are two citrus rootstocks with contrasting tolerance to drought and heat stress and have been used in this work as a model for the study of plant tolerance to the combination of drought and high temperatures. According to our results, leaf integrity and photosynthetic machinery are less affected in Carrizo than in Cleopatra under combined conditions of drought and heat stress. The pattern of accumulation of three proteins (APX, HSP101 and HSP17.6) involved in abiotic stress tolerance shows that they do not accumulate under water stress conditions individually applied. However, contents of APX and HSP101 are higher in Carrizo than in Cleopatra under stress combination whereas HSP17.6 has a similar behavior in both types of plants. This, together with a better stomatal control and a higher APX activity of Carrizo, contributes to the higher tolerance of Carrizo plants to the combination of stresses and point to it as a better rootstock than Cleopatra (traditionally used in areas with scare water supplies) under the predictable future climatic conditions with frequent periods of drought combined with high temperatures. This work also provides the basis for testing the tolerance of different citrus varieties grafted on these rootstocks and growing under different field conditions.

Regulation of citrus responses to the combined action of drought and high temperatures depends on the severity of water deprivation.

Plants grown in natural environment are regularly subjected to different combinations of abiotic stresses. Recent studies revealed that citrus plants subjected to a combination of severe drought and high temperatures displayed specific physiological, hormonal, molecular and metabolic responses. In the present study, we have performed a long-term experiment combining moderate drought and heat in Cleopatra mandarin to evaluate the impact of the stress-sequence, intensity and duration. Our results support previous observation of high sensitivity of Cleopatra mandarin to abiotic stresses that include high temperatures. In this sense, a combination of drought and heat stress negatively impacts Cleopatra seedlings independently of the drought intensity. However, some responses to combined drought and heat depend on drought intensity, especially those involved in stomatal regulation. The intricate natural environment, abiotic stress combinations and global climatic changes increase the complexity of studying plant responses to stress factors in the laboratory. Consequently, new experimental approaches taking in consideration different stress combinations should be implemented to study the viability of Cleopatra mandarin as a rootstock in a rapidly changing environment.

Plant adaptations to the combination of drought and high temperatures.

Under field conditions crops are routinely subjected to a number of different abiotic stress factors simultaneously. Recent studies revealed that the response of plants to a combination of different abiotic stresses is unique and cannot be directly extrapolated from simply studying each of the different stresses applied individually. These studies have also identified specific regulatory transcripts, combinations of metabolites and proteins, and physiological responses that are unique to specific stress combinations, highlighting the importance of studying abiotic stress combination in plants. Here we describe the interactions between drought and other abiotic stresses with emphasis on drought and heat stress. We compile new data about the different molecular, physiological and metabolic adaptations of different plants and crops to this stress combination and we highlight the importance of reactive oxygen species (ROS) metabolism and stomatal responses for plant acclimation to drought and heat stress combination. We further emphasize the need for developing crops with enhanced tolerance to drought and heat stress combination in order to mitigate the negative impacts of predicted global climatic changes on agricultural production worldwide.

Modulation of Antioxidant Defense System Is Associated with Combined Drought and Heat Stress Tolerance in Citrus.

Drought and high temperatures are two major abiotic stress factors that often occur simultaneously in nature, affecting negatively crop performance and yield. Moreover, these environmental challenges induce oxidative stress in plants through the production of reactive oxygen species (ROS). Carrizo citrange and Cleopatra mandarin are two citrus genotypes with contrasting ability to cope with the combination of drought and heat stress. In this work, a direct relationship between an increased antioxidant activity and stress tolerance is reported. According to our results, the ability of Carrizo plants to efficiently coordinate superoxide dismutase (SOD), ascorbate peroxidase (APX), catalase (CAT), and glutathione reductase (GR) activities involved in ROS detoxification along with the maintenance of a favorable GSH/GSSG ratio could be related to their relative tolerance to this stress combination. On the other hand, the increment of SOD activity and the inefficient GR activation along with the lack of CAT and APX activities in Cleopatra plants in response to the combination of drought and heat stress, could contribute to an increased oxidative stress and the higher sensibility of this citrus genotype to this stress combination.

ABA is required for the accumulation of APX1 and MBF1c during a combination of water deficit and heat stress.

Abscisic acid (ABA) plays a key role in plant acclimation to abiotic stress. Although recent studies suggested that ABA could also be important for plant acclimation to a combination of abiotic stresses, its role in this response is currently unknown. Here we studied the response of mutants impaired in ABA signalling (abi1-1) and biosynthesis (aba1-1) to a combination of water deficit and heat stress. Both mutants displayed reduced growth, biomass, and survival when subjected to stress combination. Focusing on abi1-1, we found that although its stomata had an impaired response to water deficit, remaining significantly more open than wild type, its stomatal aperture was surprisingly reduced when subjected to the stress combination. Stomatal closure during stress combination in abi1-1 was accompanied by higher levels of H2O2 in leaves, suggesting that H2O2 might play a role in this response. In contrast to the almost wild-type stomatal closure phenotype of abi1-1 during stress combination, the accumulation of ascorbate peroxidase 1 and multiprotein bridging factor 1c proteins, required for acclimation to a combination of water deficit and heat stress, was significantly reduced in abi1-1 Our findings reveal a key function for ABA in regulating the accumulation of essential proteins during a combination of water deficit and heat stress.