Stomatal pores are formed by pairs of specialized epidermal guard cells and serve as major gateways for both CO 2 influx into plants from the atmosphere and transpirational water loss of plants. Because they regulate stomatal pore apertures via integration of both endogenous hormonal stimuli and environmental signals, guard cells have been highly developed as a model system to dissect the dynamics and mechanisms of plant-cell signaling. The stress hormone ABA and elevated levels of CO 2 activate complex signaling pathways in guard cells that are mediated by kinases/phosphatases, secondary messengers, and ion channel regulation. Recent research in guard cells has led to a new hypothesis for how plants achieve specificity in intracellular calcium signaling: CO 2 and ABA enhance (prime) the calcium sensitivity of downstream calciumsignaling mechanisms. Recent progress in identification of early stomatal signaling components are reviewed here, including ABA receptors and CO 2 -binding response proteins, as well as systems approaches that advance our understanding of guard cell-signaling mechanisms.

The continuing rise in atmospheric CO2 causes closing of stomatal pores in leaves and thus globally affects CO2 influx into plants, water use efficiency and leaf heat stress1–4. However, the CO2-binding proteins that control this response remain unknown. Moreover, the cell type that responds to CO2, mesophyll or guard cells, and whether photosynthesis mediates this response are matters of debate5–8. We demonstrate that Arabidopsis double mutant plants in the β-carbonic anhydrases, βCA1 and βCA4, display impaired CO2-regulation of stomatal movements and increased stomatal density, but retain functional abscisic-acid and blue-light responses.βCA-mediated CO2-triggered stomatal movements are not, in-first-order, linked to leaf-photosynthesis and can function in guard cells. Furthermore, guard cell βCA-over-expression plants exhibit enhanced water use efficiency. Guard cell-expression of mammalian αCAII complements ca1ca4 shows that carbonic anhydrase-mediated catalysis is an important mechanism for βCA-mediated CO2-induced stomatal closing and patch clamp analyses indicate that CO2/HCO3−transfers the signal to anion channel regulation. These findings, together with ht1-29 epistasis analysis demonstrate that carbonic anhydrases function early in the CO2 signalling pathway that controls gas-exchange between plants and the atmosphere.

Summary Coordinated regulation of protection mechanisms against environmental abiotic stress and pathogen attack is essential for plant adaptation and survival. Initial abiotic stress can interfere with disease resistance signaling [1–6]. Conversely, initial plant immune signaling may interrupt subsequent ABA signal transduction [7, 8]. However, the processes involved in cross talk between these signaling networks have not been determined. By screening a 9,600 compound chemical library, we identified a small molecule DFPM that rapidly down-regulates ABA-dependent gene expression and also inhibits ABA-induced stomatal closure. Transcriptome analyses show that DFPM also stimulates expression of plant defense-related genes. Major early regulators of pathogen resistance responses, including EDS1, PAD4, RAR1, and SGT1b, are required for DFPM- and notably also for Pseudomonas-interference with ABA signal transduction, whereas salicylic acid, EDS16 and NPR1 are not necessary. While DFPM does not interfere with early ABA perception by PYR/RCAR receptors or ABA-activation of SnRK2 kinases, it disrupts cytosolic Ca2+ signaling and downstream anion channel activation in a pad4-dependent manner. Our findings provide evidence that activation of EDS1/PAD4-dependent plant immune responses rapidly disrupts ABA signal transduction and this occurs at the level of Ca2+ signaling, illuminating how the initial biotic stress pathway interferes with ABA signaling.

Guard cells regulate plant gas exchange and transpiration by modulation of stomatal aperture upon integrating external cues like photosynthetic effective illumination, CO2 levels and water availability and internal signals like abscisic acid (ABA). Being pores, stomata constitute a natural entry site for potentially harmful microbes. To prevent microbial invasion, stomata close upon perception of microbe-associated molecular patterns (MAMPs), and this represents an important layer of active immunity at the preinvasive level. The signaling pathways leading to stomatal closure triggered by biotic and abiotic stresses employ several common components, such as reactive oxygen species, calcium, kinases, and hormones, suggesting considerable intersection between MAMP- and ABA-induced stomatal closures, which we will discuss in this review.

ORCID IDs: 0000-0003-0538-6646 (H.H.); 0000-0001-6675-273X (M.B.).Elevated carbon dioxide (CO 2 ) in leaves closes stomatal apertures. Research has shown key functions of the b-carbonic anhydrases (bCA1 and bCA4) in rapid CO 2 -induced stomatal movements by catalytic transmission of the CO 2 signal in guard cells. However, the underlying mechanisms remain unclear, because initial studies indicate that these Arabidopsis (Arabidopsis thaliana) bCAs are targeted to distinct intracellular compartments upon expression in tobacco (Nicotiana benthamiana) cells. Which cellular location of these enzymes plays a key role in native guard cells in CO 2 -regulated stomatal movements remains unknown. Here, we express fluorescently tagged CAs in guard cells of ca1ca4 double-mutant plants and show that the specific locations of bCA4 at the plasma membrane and bCA1 in native guard cell chloroplasts each can mediate rapid CO 2 control of stomatal movements. Localization and complementation analyses using a mammalian aCAII-yellow fluorescent protein in guard cells further show that cytoplasmic localization is also sufficient to restore CO 2 regulation of stomatal conductance. Mathematical modeling of cellular CO 2 catalysis suggests that the dynamics of the intracellular HCO 3 2 concentration change in guard cells can be driven by plasma membrane and cytoplasmic localizations of CAs but not as clearly by chloroplast targeting. Moreover, modeling supports the notion that the intracellular HCO 3 2 concentration dynamics in guard cells are a key mechanism in mediating CO 2 -regulated stomatal movements but that an additional chloroplast role of CAs exists that has yet to be identified.