ROS are universal cell-to-cell stress signals.

The interplay between reactive oxygen species (ROS) and the redox state of cells is deeply rooted in the biology of almost all organisms, regulating development, growth, and responses to the environment. Recent studies revealed that the ROS levels and redox state of one cell can be transmitted, as an information 'state' or 'currency', to other cells and spread by cell-to-cell communication within an entire community of cells or an organism. Here, we discuss the different pathways that mediate cell-to-cell signaling in plants, their hierarchy, and the different mechanisms that transmit ROS/redox signaling between different cells. We further hypothesize that ROS/redox signaling between different organisms could play a key role within the 'one world' principle, impacting human health and our future.

Functional analysis of reactive oxygen species-driven stress systemic signalling, interplay and acclimation.

Reactive oxygen species (ROS) play a critical role in plant development and stress responses, acting as key components in rapid signalling pathways. The 'ROS wave' triggers essential acclimation processes, ultimately ensuring plant survival under diverse challenges. This review explores recent advances in understanding the composition and functionality of the ROS wave within plant cells. During their initiation and propagation, ROS waves interact with other rapid signalling pathways, hormones and various molecular compounds. Recent research sheds light on the intriguing lack of a rigid hierarchy governing these interactions, highlighting a complex interplay between diverse signals. Notably, ROS waves culminate in systemic acclimation, a crucial outcome for enhanced stress tolerance. This review emphasizes the versatility of ROS, which act as flexible players within a network of short- and long-term factors contributing to plant stress resilience. Unveiling the intricacies of these interactions between ROS and various signalling molecules holds immense potential for developing strategies to augment plant stress tolerance, contributing to improved agricultural practices and overall ecosystem well-being.

Propagation of a rapid cell-to-cell H2O2 signal over long distances in a monolayer of cardiomyocyte cells.

Cell-to-cell communication plays a cardinal role in the biology of multicellular organisms. H2O2 is an important cell-to-cell signaling molecule involved in the response of mammalian cells to wounding and other stimuli. We previously identified a signaling pathway that transmits wound-induced cell-to-cell H2O2 signals within minutes over long distances, measured in centimeters, in a monolayer of cardiomyocytes. Here we report that this long-distance H2O2 signaling pathway is accompanied by enhanced accumulation of cytosolic H2O2 and altered redox state in cells along its path. We further show that it requires the production of superoxide, as well as the function of gap junctions, and that it is accompanied by changes in the abundance of hundreds of proteins in cells along its path. Our findings highlight the existence of a unique and rapid long-distance H2O2 signaling pathway that could play an important role in different inflammatory responses, wound responses/healing, cardiovascular disease, and/or other conditions.

ROS are evolutionary conserved cell-to-cell stress signals.

Cell-to-cell communication is fundamental to multicellular organisms and unicellular organisms living in a microbiome. It is thought to have evolved as a stress- or quorum-sensing mechanism in unicellular organisms. A unique cell-to-cell communication mechanism that uses reactive oxygen species (ROS) as a signal (termed the "ROS wave") was identified in flowering plants. This process is essential for systemic signaling and plant acclimation to stress and can spread from a small group of cells to the entire plant within minutes. Whether a similar signaling process is found in other organisms is however unknown. Here, we report that the ROS wave can be found in unicellular algae, amoeba, ferns, mosses, mammalian cells, and isolated hearts. We further show that this process can be triggered in unicellular and multicellular organisms by a local stress or H2O2 treatment and blocked by the application of catalase or NADPH oxidase inhibitors and that in unicellular algae it communicates important stress-response signals between cells. Taken together, our findings suggest that an active process of cell-to-cell ROS signaling, like the ROS wave, evolved before unicellular and multicellular organisms diverged. This mechanism could have communicated an environmental stress signal between cells and coordinated the acclimation response of many different cells living in a community. The finding of a signaling process, like the ROS wave, in mammalian cells further contributes to our understanding of different diseases and could impact the development of drugs that target for example cancer or heart disease.

Phytochrome B regulates reactive oxygen signaling during abiotic and biotic stress in plants.

Reactive oxygen species (ROS) and the photoreceptor protein phytochrome B (phyB) play a key role in plant acclimation to stress. However, how phyB that primarily functions in the nuclei impacts ROS signaling mediated by respiratory burst oxidase homolog (RBOH) proteins that reside on the plasma membrane, during stress, is unknown. Arabidopsis thaliana and Oryza sativa mutants, RNA-Seq, bioinformatics, biochemistry, molecular biology, and whole-plant ROS imaging were used to address this question. Here, we reveal that phyB and RBOHs function as part of a key regulatory module that controls apoplastic ROS production, stress-response transcript expression, and plant acclimation in response to excess light stress. We further show that phyB can regulate ROS production during stress even if it is restricted to the cytosol and that phyB, respiratory burst oxidase protein D (RBOHD), and respiratory burst oxidase protein F (RBOHF) coregulate thousands of transcripts in response to light stress. Surprisingly, we found that phyB is also required for ROS accumulation in response to heat, wounding, cold, and bacterial infection. Our findings reveal that phyB plays a canonical role in plant responses to biotic and abiotic stresses, regulating apoplastic ROS production, possibly while at the cytosol, and that phyB and RBOHD/RBOHF function in the same regulatory pathway.

Jasmonic acid and salicylic acid modulate systemic reactive oxygen species signaling during stress responses.

Plants can send long-distance cell-to-cell signals from a single tissue subjected to stress to the entire plant. This ability is termed "systemic signaling" and is essential for plant acclimation to stress and/or defense against pathogens. Several signaling mechanisms are associated with systemic signaling, including the reactive oxygen species (ROS) wave, calcium wave, hydraulic wave, and electric signals. The ROS wave coordinates multiple physiological, molecular, and metabolic responses among different parts of the plant and is essential for systemic acquired acclimation (SAA) to stress. In addition, it is linked with several plant hormones, including jasmonic acid (JA), salicylic acid (SA), and abscisic acid (ABA). However, how these plant hormones modulate the ROS wave and whether they are required for SAA is not clear. Here we report that SA and JA play antagonistic roles in modulating the ROS wave in Arabidopsis (Arabidopsis thaliana). While SA augments the ROS wave, JA suppresses it during responses to local wounding or high light (HL) stress treatments. We further show that ethylene and ABA are essential for regulation of the ROS wave during systemic responses to local wounding treatment. Interestingly, we found that the redox-response protein NONEXPRESSOR OF PATHOGENESIS RELATED PROTEIN 1 is required for systemic ROS accumulation in response to wounding or HL stress, as well as for SAA to HL stress. Taken together, our findings suggest that interplay between JA and SA might regulate systemic signaling and SAA during responses of plants to abiotic stress or wounding.

Propagation of a rapid cell-to-cell H 2 O 2 signal over long distances in a monolayer of cardiomyocyte cells.

Cell-to-cell communication plays a cardinal role in the biology of multicellular organisms. H 2 O 2 is an important cell-to-cell signaling molecule involved in the response of mammalian cells to wounding and other stimuli. We previously identified a signaling pathway that transmits wound-induced cell-to-cell H 2 O 2 signals within minutes over long distances, measured in centimeters, in a monolayer of cardiomyocytes. Here we report that this long-distance H 2 O 2 signaling pathway is accompanied by enhanced accumulation of cytosolic H 2 O 2 and altered redox state in cells along its path. We further show that it requires the production of superoxide, as well as the function of gap junctions, and that it is accompanied by changes in the abundance of hundreds of proteins in cells along its path. Our findings highlight the existence of a unique and rapid long-distance H 2 O 2 signaling pathway that could play an important role in different inflammatory responses, wound responses/healing, cardiovascular disease, and/or other conditions. Highlights: Wounding induces an H 2 O 2 cell-to-cell signal in a monolayer of cardiomyocytes. The cell-to-cell signal requires H 2 O 2 and O 2 · - accumulation along its path. The signal propagates over several centimeters changing the redox state of cells.Changes in the abundance of hundreds of proteins accompanies the signal.The cell-to-cell signal requires paracrine and juxtacrine signaling.

ROS and redox regulation of cell-to-cell and systemic signaling in plants during stress.

Stress results in the enhanced accumulation of reactive oxygen species (ROS) in plants, altering the redox state of cells and triggering the activation of multiple defense and acclimation mechanisms. In addition to activating ROS and redox responses in tissues that are directly subjected to stress (termed 'local' tissues), the sensing of stress in plants triggers different systemic signals that travel to other parts of the plant (termed 'systemic' tissues) and activate acclimation and defense mechanisms in them; even before they are subjected to stress. Among the different systemic signals triggered by stress in plants are electric, calcium, ROS, and redox waves that are mobilized in a cell-to-cell fashion from local to systemic tissues over long distances, sometimes at speeds of up to several millimeters per second. Here, we discuss new studies that identified various molecular mechanisms and proteins involved in mediating systemic signals in plants. In addition, we highlight recent studies that are beginning to unravel the mode of integration and hierarchy of the different systemic signals and underline open questions that require further attention. Unraveling the role of ROS and redox in plant stress responses is highly important for the development of climate resilient crops.

Multi-omic characterization of bifunctional peroxidase 4-coumarate 3-hydroxylase knockdown in Brachypodium distachyon provides insights into lignin modification-associated pleiotropic effects.

A bifunctional peroxidase enzyme, 4-coumarate 3-hydroxylase (C3H/APX), provides a parallel route to the shikimate shunt pathway for the conversion of 4-coumarate to caffeate in the early steps of lignin biosynthesis. Knockdown of C3H/APX (C3H/APX-KD) expression has been shown to reduce the lignin content in Brachypodium distachyon. However, like many other lignin-modified plants, C3H/APX-KDs show unpredictable pleiotropic phenotypes, including stunted growth, delayed senescence, and reduced seed yield. A system-wide level understanding of altered biological processes in lignin-modified plants can help pinpoint the lignin-modification associated growth defects to benefit future studies aiming to negate the yield penalty. Here, a multi-omic approach was used to characterize molecular changes resulting from C3H/APX-KD associated lignin modification and negative growth phenotype in Brachypodium distachyon. Our findings demonstrate that C3H/APX knockdown in Brachypodium stems substantially alters the abundance of enzymes implicated in the phenylpropanoid biosynthetic pathway and disrupt cellular redox homeostasis. Moreover, it elicits plant defense responses associated with intracellular kinases and phytohormone-based signaling to facilitate growth-defense trade-offs. A deeper understanding along with potential targets to mitigate the pleiotropic phenotypes identified in this study could aid to increase the economic feasibility of lignocellulosic biofuel production.

HPCA1 is required for systemic reactive oxygen species and calcium cell-to-cell signaling and plant acclimation to stress.

Reactive oxygen species (ROS), produced by respiratory burst oxidase homologs (RBOHs) at the apoplast, play a key role in local and systemic cell-to-cell signaling, required for plant acclimation to stress. Here we reveal that the Arabidopsis thaliana leucine-rich-repeat receptor-like kinase H2O2-INDUCED CA2+ INCREASES 1 (HPCA1) acts as a central ROS receptor required for the propagation of cell-to-cell ROS signals, systemic signaling in response to different biotic and abiotic stresses, stress responses at the local and systemic tissues, and plant acclimation to stress, following a local treatment of high light (HL) stress. We further report that HPCA1 is required for systemic calcium signals, but not systemic membrane depolarization responses, and identify the calcium-permeable channel MECHANOSENSITIVE ION CHANNEL LIKE 3, CALCINEURIN B-LIKE CALCIUM SENSOR 4 (CBL4), CBL4-INTERACTING PROTEIN KINASE 26 and Sucrose-non-fermenting-1-related Protein Kinase 2.6/OPEN STOMATA 1 (OST1) as required for the propagation of cell-to-cell ROS signals. In addition, we identify serine residues S343 and S347 of RBOHD (the putative targets of OST1) as playing a key role in cell-to-cell ROS signaling in response to a local application of HL stress. Our findings reveal that HPCA1 plays a key role in mediating and coordinating systemic cell-to-cell ROS and calcium signals required for plant acclimation to stress.

Reactive oxygen species signalling in plant stress responses.

Reactive oxygen species (ROS) are key signalling molecules that enable cells to rapidly respond to different stimuli. In plants, ROS play a crucial role in abiotic and biotic stress sensing, integration of different environmental signals and activation of stress-response networks, thus contributing to the establishment of defence mechanisms and plant resilience. Recent advances in the study of ROS signalling in plants include the identification of ROS receptors and key regulatory hubs that connect ROS signalling with other important stress-response signal transduction pathways and hormones, as well as new roles for ROS in organelle-to-organelle and cell-to-cell signalling. Our understanding of how ROS are regulated in cells by balancing production, scavenging and transport has also increased. In this Review, we discuss these promising developments and how they might be used to increase plant resilience to environmental stress.

Aboveground plant-to-plant electrical signaling mediates network acquired acclimation.

Systemic acquired acclimation and wound signaling require the transmission of electrical, calcium, and reactive oxygen species (ROS) signals between local and systemic tissues of the same plant. However, whether such signals can be transmitted between two different plants is largely unknown. Here, we reveal a new type of plant-to-plant aboveground direct communication involving electrical signaling detected at the surface of leaves, ROS, and photosystem networks. A foliar electrical signal induced by wounding or high light stress applied to a single dandelion leaf can be transmitted to a neighboring plant that is in direct contact with the stimulated plant, resulting in systemic photosynthetic, oxidative, molecular, and physiological changes in both plants. Furthermore, similar aboveground changes can be induced in a network of plants serially connected via touch. Such signals can also induce responses even if the neighboring plant is from a different plant species. Our study demonstrates that electrical signals can function as a communication link between transmitter and receiver plants that are organized as a network (community) of plants. This process can be described as network-acquired acclimation.

Differential regulation of flower transpiration during abiotic stress in annual plants.

Heat waves occurring during droughts can have a devastating impact on yield, especially if they happen during the flowering and seed set stages of the crop cycle. Global warming and climate change are driving an alarming increase in the frequency and intensity of combined drought and heat stress episodes, critically threatening global food security. Because high temperature is detrimental to reproductive processes, essential for plant yield, we measured the inner temperature, transpiration, sepal stomatal aperture, hormone concentrations and transcriptomic response of closed soybean flowers developing on plants subjected to a combination of drought and heat stress. Here, we report that, during a combination of drought and heat stress, soybean plants prioritize transpiration through flowers over transpiration through leaves by opening their flower stomata, while keeping their leaf stomata closed. This acclimation strategy, termed 'differential transpiration', lowers flower inner temperature by about 2-3°C, protecting reproductive processes at the expense of vegetative tissues. Manipulating stomatal regulation, stomatal size and/or stomatal density of flowers could serve as a viable strategy to enhance the yield of different crops and mitigate some of the current and future impacts of global warming and climate change on agriculture.

Extracellular ATP plays an important role in systemic wound response activation.

Mechanical wounding occurs in plants during biotic or abiotic stresses and is associated with the activation of long-distance signaling pathways that trigger wound responses in systemic tissues. Among the different systemic signals activated by wounding are electric signals, calcium, hydraulic, and reactive oxygen species (ROS) waves. The release of glutamate (Glu) from cells at the wounded tissues was recently proposed to trigger systemic signal transduction pathways via GLU-LIKE RECEPTORs (GLRs). However, the role of another important compound released from cells during wounding (extracellular ATP [eATP]) in triggering systemic responses is not clear. Here, we show in Arabidopsis (Arabidopsis thaliana) that wounding results in the accumulation of nanomolar levels of eATP and that these levels are sufficient to trigger the systemic ROS wave. We further show that the triggering of the ROS wave by eATP during wounding requires the PURINORECEPTOR 2 KINASE (P2K) receptor. Application of eATP to unwounded leaves triggered the ROS wave, and the activation of the ROS wave by wounding or eATP application was suppressed in mutants deficient in P2Ks (e.g. p2k1-3, p2k2, and p2k1-3p2k2). In addition, expression of systemic wound response (SWR) transcripts was suppressed in mutants deficient in P2Ks during wounding. Interestingly, the effect of Glu and eATP application on ROS wave activation was not additive, suggesting that these two compounds function in the same pathway to trigger the ROS wave. Our findings reveal that in addition to sensing Glu via GLRs, eATP sensed by P2Ks plays a key role in the triggering of SWRs in plants.

Cadmium interference with iron sensing reveals transcriptional programs sensitive and insensitive to reactive oxygen species.

Iron (Fe) is an essential micronutrient whose uptake is tightly regulated to prevent either deficiency or toxicity. Cadmium (Cd) is a non-essential element that induces both Fe deficiency and toxicity; however, the mechanisms behind these Fe/Cd-induced responses are still elusive. Here we explored Cd- and Fe-associated responses in wild-type Arabidopsis and in a mutant that overaccumulates Fe (opt3-2). Gene expression profiling revealed a large overlap between transcripts induced by Fe deficiency and Cd exposure. Interestingly, the use of opt3-2 allowed us to identify additional gene clusters originally induced by Cd in the wild type but repressed in the opt3-2 background. Based on the high levels of H2O2 found in opt3-2, we propose a model where reactive oxygen species prevent the induction of genes that are induced in the wild type by either Fe deficiency or Cd. Interestingly, a defined cluster of Fe-responsive genes was found to be insensitive to this negative feedback, suggesting that their induction by Cd is more likely to be the result of an impaired Fe sensing. Overall, our data suggest that Fe deficiency responses are governed by multiple inputs and that a hierarchical regulation of Fe homeostasis prevents the induction of specific networks when Fe and H2O2 levels are elevated.

Combination of Antioxidant Enzyme Overexpression and N-Acetylcysteine Treatment Enhances the Survival of Bone Marrow Mesenchymal Stromal Cells in Ischemic Limb in Mice With Type 2 Diabetes.

Background Therapy with mesenchymal stem cells remains a promising but challenging approach to critical limb ischemia in diabetes because of the dismal cell survival. Methods and Results Critical limb ischemia in type 2 diabetes mouse model was used to explore the impact of diabetic limb ischemia on the survival of bone marrow mesenchymal stromal cells (bMSCs). Inhibition of intracellular reactive oxygen species was achieved with concomitant overexpression of superoxide dismutase (SOD)-1 and glutathione peroxidase-1 in the transplanted bMSCs, and extracellular reactive oxygen species was attenuated using SOD-3 overexpression and N-acetylcysteine treatment. In vivo optical fluorescence imaging and laser Doppler perfusion imaging were used to track cell retention and determine blood flow in diabetic ischemic limb, respectively. Survival of the transplanted bMSCs was significantly decreased in diabetic ischemic limb compared with the control. In vitro study indicated that advanced glycation end products, not high glucose, significantly decreased the proliferation of bMSCs and increased their apoptosis associated with increased reactive oxygen species production and selective reduction of SOD-1 and SOD-3. In vivo study demonstrated that concomitant overexpression of SOD-1, SOD-3, and glutathione peroxidase-1, or host treatment with N-acetylcysteine, significantly enhanced in vivo survival of transplanted bMSCs, and improved critical limb ischemia in diabetic mice. Combination of triple antioxidant enzyme overexpression in bMSCs with host N-acetylcysteine treatment further improved bMSC survival with enhanced circulatory and functional recovery from diabetic critical limb ischemia. Conclusions Simultaneous suppression of reactive oxygen species from transplanted bMSCs and host tissue could additively enhance bMSC survival in diabetic ischemic limb with increased therapeutic efficacy in diabetes.

Integration of electric, calcium, reactive oxygen species and hydraulic signals during rapid systemic signaling in plants.

The sensing of abiotic stress, mechanical injury or pathogen attack by a single plant tissue results in the activation of systemic signals that travel from the affected tissue to the entire plant. This process is essential for plant survival during stress and is termed systemic signaling. Among the different signals triggered during this process are calcium, electric, reactive oxygen species and hydraulic signals. These are thought to propagate at rapid rates through the plant vascular bundles and to regulate many of the systemic processes essential for plant survival. Although the different signals activated during systemic signaling are thought to be interlinked, their coordination and hierarchy still need to be determined. Here, using a combination of advanced whole-plant imaging and hydraulic pressure measurements, we studied the activation of all four systemic signals in wild-type and different Arabidopsis thaliana mutants subjected to a local treatment of high-light (HL) stress or wounding. Our findings reveal that activation of systemic membrane potential, calcium, reactive oxygen species and hydraulic pressure signals, in response to wounding, is dependent on glutamate receptor-like proteins 3.3 and 3.6. In contrast, in response to HL stress, systemic changes in calcium and membrane potential depended on glutamate receptor-like 3.3 and 3.6, while systemic hydraulic signals did not. We further show that plasmodesmata functions are required for systemic changes in membrane potential and calcium during responses to HL stress or wounding. Our findings shed new light on the different mechanisms that integrate different systemic signals in plants during stress.

Plasmodesmata-localized proteins and ROS orchestrate light-induced rapid systemic signaling in Arabidopsis.

Systemic signaling and systemic acquired acclimation (SAA) are key to the survival of plants during episodes of abiotic stress. These processes depend on a continuous chain of cell-to-cell signaling events that extends from the initial tissue that senses the stress (the local tissue) to the entire plant (systemic tissues). Reactive oxygen species (ROS) and Ca2+ are key signaling molecules thought to be involved in this cell-to-cell mechanism. Here, we report that the systemic response of Arabidopsis thaliana to a local treatment of high light stress, which resulted in local ROS accumulation, required ROS generated by respiratory burst oxidase homolog D (RBOHD). ROS increased cell-to-cell transport and plasmodesmata (PD) pore size in a manner dependent on PD-localized protein 1 (PDLP1) and PDLP5, and this process was required for the propagation of the systemic ROS signals and SAA. Furthermore, aquaporins and several Ca2+-permeable channels in the glutamate receptor-like (GLR), mechanosensitive small conductance-like (MSL), and cyclic nucleotide-gated (CNGC) families were involved in this systemic signaling process. However, we determined that these channels were required primarily to amplify the systemic signal in each cell along the path of the systemic ROS wave, as well as to establish local and systemic acclimation. Thus, PD and RBOHD-generated ROS orchestrate light stress-induced rapid cell-to-cell spread of systemic signals in Arabidopsis.

Integration of reactive oxygen species and hormone signaling during abiotic stress.

Each year, abiotic stress conditions such as drought, heat, salinity, cold and particularly their different combinations, inflict a heavy toll on crop productivity worldwide. The effects of these adverse conditions on plant productivity are becoming ever more alarming in recent years in light of the increased rate and intensity of global climatic changes. Improving crop tolerance to abiotic stress conditions requires a deep understanding of the response of plants to changes in their environment. This response is dependent on early and late signal transduction events that involve important signaling molecules such as reactive oxygen species (ROS), different plant hormones and other signaling molecules. It is the integration of these signaling events, mediated by an interplay between ROS and different plant hormones that orchestrates the plant response to abiotic stress and drive changes in transcriptomic, metabolic and proteomic networks that lead to plant acclimation and survival. Here we review some of the different studies that address hormone and ROS integration during the response of plants to abiotic stress. We further highlight the integration of ROS and hormone signaling during early and late phases of the plant response to abiotic stress, the key role of respiratory burst oxidase homologs in the integration of ROS and hormone signaling during these phases, and the involvement of hormone and ROS in systemic signaling events that lead to systemic acquired acclimation. Lastly, we underscore the need to understand the complex interactions that occur between ROS and different plant hormones during stress combinations.