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Chloroplast-to-nucleus retrograde signaling is essential for cell function, acclimation to fluctuating environmental conditions, plant growth and development. The vast majority of chloroplast proteins are nuclear-encoded, and must be imported into the organelle after synthesis in the cytoplasm. This import is essential for the development of fully functional chloroplasts. On the other hand, functional chloroplasts act as sensors of environmental changes and can trigger acclimatory responses that influence nuclear gene expression. Signaling via mobile transcription factors (TFs) has been recently recognized as a way of communication between organelles and the nucleus. In this study, we performed a targeted reverse genetic screen to identify dual-localized TFs involved in chloroplast retrograde signaling during stress responses. We found that CHLOROPLAST IMPORT APPARATUS 2 (CIA2) has a functional plastid transit peptide, and can be located both in chloroplasts and the nucleus. Further, we found that CIA2, along with its homolog CIA2-like (CIL) are involved in the regulation of Arabidopsis responses to UV-AB, high light and heat shock. Finally, our results suggest that both CIA2 and CIL are crucial for chloroplast translation. Our results contribute to a deeper understanding of signaling events in the chloroplast-nucleus cross-talk.

Guard cells regulate plant gas exchange by controlling the aperture of stomatal pores. The process of stomatal closure involves a multi-input signaling network that governs the activity of ion channels, which in turn regulate guard cell turgor pressure and volume. Here we describe a forward genetic screen to identify novel components involved in stomatal movements. Through an ozone-sensitivity approach combined with whole-rosette gas exchange analysis, 130 mutants of established stomatal regulators and 76 novel mutants impaired in stomatal closure were identified. One of the novel mutants was mapped to MURUS1 (MUR1), the first enzyme in de novo GDP-L-fucose biosynthesis. Defects in synthesis or import of GDP-L-Fuc into the Golgi apparatus resulted in impaired stomatal closure to multiple stimuli. Stomatal phenotypes observed in mur1 were independent from the canonical guard cell signaling and instead could be related to altered mechanical properties of guard cell walls. Impaired fucosylation of xyloglucan, N-linked glycans and arabinogalactan proteins did not explain the aberrant function of mur1 stomata, however our data suggest that the stomatal phenotypes observed in mur1 can at least partially be attributed to defective dimerization of rhamnogalactouronan-II. In addition to providing the genetic framework for future studies on guard cell signaling, our work emphasizes the impact of fucose metabolism on stomatal movement.

Climate change results in increased intensity and frequency of extreme abiotic and biotic stress events. In plants, reactive oxygen species (ROS) accumulate in proportion to the level of stress and are major signalling and regulatory metabolites coordinating growth, defence, acclimation and cell death. Our knowledge of ROS homeostasis, sensing, and signalling is therefore key to understanding the impacts of climate change at the molecular level. Current research is uncovering new insights into temporal-spatial, cell-to-cell and systemic ROS signalling pathways, particularly how these affect plant growth, defence, and more recently acclimation mechanisms behind stress priming and long term stress memory. Understanding the stabilising and destabilising factors of ROS homeostasis and signalling in plants exposed to extreme and fluctuating stress will concomitantly reveal how to address future climate change challenges in global food security and biodiversity management.

Guard cells form stomatal pores that optimize photosynthetic carbon dioxide uptake with minimal water loss. Stomatal movements are controlled by complex signaling networks that respond to environmental and endogenous signals. Regulation of stomatal aperture requires coordinated activity of reactive oxygen species (ROS)-generating enzymes, signaling proteins, and downstream executors such as ion pumps, transporters, and plasma membrane channels that control guard cell turgor pressure. Accumulation of ROS in the apoplast and chloroplasts is among the earliest hallmarks of stomatal closure. Subsequent increase in cytoplasmic Ca(2+) concentration governs the activity of multiple kinases that regulate the activity of ROS-producing enzymes and ion channels. In parallel, ROS directly regulate the activity of multiple proteins via oxidative posttranslational modifications to fine-tune guard cell signaling. In this review, we summarize recent advances in the role of ROS in stomatal closure and discuss the importance of ROS in regulation of signal amplification and specificity in guard cells.

Strigolactones are a group of phytohormones that control shoot branching inArabidopsis thaliana. However, in recent years they have been shown to affect many other plant processes. We previously showed that the strigolactone perception mutantmore axillary branches 2 (max2)has increased susceptibility to plant pathogenic bacteria as a result of more open stomata as well as alterations in hormonal signalling. Here we show that both, strigolactone biosynthesis- (max3andmax4), and perception mutants (max2anddwarf14) are significantly more sensitive toPseudomonas syringaeDC3000. Moreover, in response toP. syringaeinfection, high levels of SA accumulated inmax2and this mutant was ozone sensitive. To search for the mechanisms that could explain pathogen- and ozone sensitivity we performed gene expression analysis and several different assays that explore the function of guard cells and regulation of guard cell signalling.Treatments with GR24 (a strigolactone analogue) resulted in very modest changes in defence-related gene expression. In contrast, guard cell function was clearly impaired inmax2and depending on the assay used, also inmax3, max4andd14mutants. Moreover, stomatal responses to stimuli that cause stomatal closure in wild-type plants (darkness, high CO2and ABA) were analysed in the strigolactone mutants. In darkness both strigolactone biosynthesis and perception mutants showed reduced stomatal closure, whereas the response to high CO2was impaired only inmax2andd14. The response to ABA was not impaired in any of the mutants. To position the role of MAX2 in the guard cell signalling network,max2was crossed with mutants defective in ABA biosynthesis (aba2), in guard cell ABA signalling (ost1) and a scaffold protein required for proper ion channel activity (ghr1). The stomatal conductance of double mutants was consistently higher than the corresponding single mutants, suggesting that MAX2 acts in a signalling pathway that functions in parallel to the well characterized guard cell ABA signalling pathway. We propose that the impaired defence responses ofmax2is related to more open stomata that allows increased entry of bacteria or air pollutants like ozone. Furthermore, as MAX2 appears to act in a specific branch of guard cell signalling (related to CO2signalling), this protein could be one of the elusive components that allow guard cells to distinguish between different environmental conditions.

As fixed organisms, plants are especially affected by changes in their environment and have consequently evolved extensive mechanisms for acclimation and adaptation. Initially considered by-products from aerobic metabolism, reactive oxygen species (ROS) have emerged as major regulatory molecules in plants and their roles in early signaling events initiated by cellular metabolic perturbation and environmental stimuli are now established. Here, we review recent advances in ROS signaling. Compartment-specific and cross-compartmental signaling pathways initiated by the presence of ROS are discussed. Special attention is dedicated to established and hypothetical ROS-sensing events. The roles of ROS in long-distance signaling, immune responses, and plant development are evaluated. Finally, we outline the most challenging contemporary questions in the field of plant ROS biology and the need to further elucidate mechanisms allowing sensing, signaling specificity, and coordination of multiple signals.

In plants, fluctuation of the redox balance by altered levels of reactive oxygen species (ROS) can affect many aspects of cellular physiology. ROS homeostasis is governed by a diversified set of antioxidant systems. Perturbation of this homeostasis leads to transient or permanent changes in the redox status and is exploited by plants in different stress signalling mechanisms. Understanding how plants sense ROS and transduce these stimuli into downstream biological responses is still a major challenge. ROS can provoke reversible and irreversible modifications to proteins that act in diverse signalling pathways. These oxidative post-translational modifications (Ox-PTMs) lead to oxidative damage and/or trigger structural alterations in these target proteins. Characterization of the effect of individual Ox-PTMs on individual proteins is the key to a better understanding of how cells interpret the oxidative signals that arise from developmental cues and stress conditions. This review focuses on ROS-mediated Ox-PTMs on cysteine (Cys) residues. The Cys side chain, with its high nucleophilic capacity, appears to be the principle target of ROS. Ox-PTMs on Cys residues participate in various signalling cascades initiated by plant stress hormones. We review the mechanistic aspects and functional consequences of Cys Ox-PTMs on specific target proteins in view of stress signalling events.

In plants, receptor-like kinases (RLKs) and extracellular reactive oxygen species (ROS) contribute to the communication between the environment and the interior of the cell. Apoplastic ROS production is a frequent result of RLK signaling in a multitude of cellular processes; thus, by their nature, these two signaling components are inherently linked. However, it is as yet unclear how ROS signaling downstream of receptor activation is executed. In this review, we provide a broad view of the intricate connections between RLKs and ROS signaling and describe the regulatory events that control and coordinate extracellular ROS production. We propose that concurrent initiation of ROS-dependent and -independent signaling linked to RLKs might be a critical element in establishing cellular responses. Furthermore, we discuss the possible ROS sensing mechanisms in the context of the biochemical environment in the apoplast. We suggest that RLK-dependent modulation of apoplastic and intracellular conditions facilitates ROS perception and signaling. Based on data from plant and animal models, we argue that specific RLKs could be components of the ROS sensing machinery or ROS sensors. The importance of the crosstalk between RLK and ROS signaling is discussed in the context of stomatal immunity. Finally, we highlight challenges in the understanding of these signaling processes and provide perspectives for future research.

The nuclear male sterility gene ms8 is expected to facilitate the production of sweet pepper (Capsicum annuum L.) hybrids as it provides means for hybridization without the labor-intensive hand emasculation of female inbred lines. The development of molecular markers linked to ms8 locus will help the breeding practice for the selection of hybrid parental lines. In this study, F2 population resulting from a cross between the sweet pepper male sterile line 320 and the male fertile variety Elf was used to identify DNA markers linked to the ms8 locus. With the use of RAPD–BSA technique, seven markers linked to the ms8 locus were found. Four of them were converted into SCAR markers. In addition, two COSII/CAPS markers linked to the ms8 locus were identified. Comparative mapping with reference pepper maps indicated that the ms8 locus is located on the lower arm of the pepper chromosome P4. Identified markers are useful for molecular breeding, however, at present markers tightly linked to ms8 locus are still lacking. Identification of molecular markers linked to the ms8 locus and determination of its chromosomal localization are useful for fine mapping and also provide the perspective for ms8 gene cloning.

The high metabolic flux through photorespiration constitutes a significant part of the carbon cycle. Although the major enzymatic steps of the photorespiratory pathway are well characterized, little information is available on the functional significance of photorespiration beyond carbon recycling. Particularly important in this respect is the peroxisomal catalase activity which removes photorespiratory H 2 O 2 generated during the oxidation of glycolate to glyoxylate, thus maintaining the cellular redox homeostasis governing the perception, integration and execution of stress responses. We have previously performed a chemical screen that led to the identification of 34 small molecules that alleviate the negative effects of photorespiration in Arabidopsis thaliana mutants lacking peroxisomal catalase ( cat2 ). In parallel, we screened a mutagenized cat2 population and looked for second-site mutations that similarly attenuate the effects of photorespiratory conditions in cat2 plants. Currently, we characterize putative protein targets of some of the chemical hits and investigate the fine-tuning roles of the confirmed mutations on the photorespiratory phenotype of cat2 .

Reactive oxygen species (ROS) have been shown to be potent signaling molecules. Today, oxidation of cysteine residues is a well-recognized posttranslational protein modification, but the signaling processes steered by such oxidations are poorly understood. To gain insight into the cysteine thiol-dependent ROS signaling in Arabidopsis thaliana, we identified the hydrogen peroxide (H2O2)-dependent sulfenome: that is, proteins with at least one cysteine thiol oxidized to a sulfenic acid. By means of a genetic construct consisting of a fusion between the C-terminal domain of the yeast (Saccharomyces cerevisiae) AP-1-like (YAP1) transcription factor and a tandem affinity purification tag, we detected ∼ 100 sulfenylated proteins in Arabidopsis cell suspensions exposed to H2O2 stress. The in vivo YAP1-based trapping of sulfenylated proteins was validated by a targeted in vitro analysis of dehydroascorbate reductase2 (DHAR2). In DHAR2, the active site nucleophilic cysteine is regulated through a sulfenic acid-dependent switch, leading to S-glutathionylation, a protein modification that protects the protein against oxidative damage.