New insights into the influence of NHX-type Cation/H+ antiporter on flower color in Phalaenopsis orchids.

Vacuolar sodium/proton Na+(K+)/H+ exchanger (NHX) influence color formation because of their effects on cellular pH and Na+/K+ homeostasis. Research regarding NHXs has mainly focused on the vacuolar NHX family members. However, the NHX functions related to Phalaenopsis flower coloration remain relatively uncharacterized. In this study, we cloned and characterized PeNHX1, a vacuolar cation/H+ antiporter-encoding gene that is highly expressed in the Phalaenopsis equestris (orchid) flower lip. Phylogenetic and sequence analyses showed that PeNHX1 is a vacuolar NHX protein family member that is similar to other known vacuolar antiporters. The PeNHX1-GFP fusion protein was clearly localized to the vacuolar membrane in a transient transfection assay. A quantitative real-time PCR analysis revealed the increased expression of PeNHX1 in different flower developmental stages. Moreover, it was more highly expressed in the lip than in the other flower organs. On the basis of virus-induced gene silencing, we determined that decreased PeNHX1 expression significantly reduces P. equestris petal coloration. Furthermore, the overexpression of PeNHX1 in Phalaenopsis Big Chili caused the pH to increase and the petal color to change from red to blue. The results indicate that NHX1 may mediates the Na + or K+/H+ exchange, thereby regulating the vacuolar pH to promote blue coloration. This research provides a theoretical basis for the development of orchid varieties with blue flowers.

Electrochemical potential enables dormant spores to integrate environmental signals.

The dormant state of bacterial spores is generally thought to be devoid of biological activity. We show that despite continued dormancy, spores can integrate environmental signals over time through a preexisting electrochemical potential. Specifically, we studied thousands of individual Bacillus subtilis spores that remain dormant when exposed to transient nutrient pulses. Guided by a mathematical model of bacterial electrophysiology, we modulated the decision to exit dormancy by genetically and chemically targeting potassium ion flux. We confirmed that short nutrient pulses result in step-like changes in the electrochemical potential of persistent spores. During dormancy, spores thus gradually release their stored electrochemical potential to integrate extracellular information over time. These findings reveal a decision-making mechanism that operates in physiologically inactive cells.

Ethylene-induced potassium transporter AcKUP2 gene is involved in kiwifruit postharvest ripening.

BACKGROUND: Potassium (K) is important in the regulation of plant growth and development. It is the most abundant mineral element in kiwifruit, and its content increases during fruit ripening. However, how K+ transporter works in kiwifruit postharvest maturation is not yet clear. RESULTS: Here, 12 K+ transporter KT/HAK/KUP genes, AcKUP1 ~ AcKUP12, were isolated from kiwifruit, and their phylogeny, genomic structure, chromosomal location, protein properties, conserved motifs and cis-acting elements were analysed. Transcription analysis revealed that AcKUP2 expression increased rapidly and was maintained at a high level during postharvest maturation, consistent with the trend of K content; AcKUP2 expression was induced by ethylene, suggesting that AcKUP2 might play a role in ripening. Fluorescence microscopy showed that AcKUP2 is localised in the plasma membrane. Cis-elements, including DER or ethylene response element (ERE) responsive to ethylene, were found in the AcKUP2 promoter sequence, and ethylene significantly enhanced the AcKUP2 promoter activity. Furthermore, we verified that AcERF15, an ethylene response factor, directly binds to the AcKUP2 promoter to promote its expression. Thus, AcKUP2 may be an important potassium transporter gene which involved in ethylene-regulated kiwifruit postharvest ripening. CONCLUSIONS: Therefore, our study establishes the first genome-wide analysis of the kiwifruit KT/HAK/KUP gene family and provides valuable information for understanding the function of the KT/HAK/KUP genes in kiwifruit postharvest ripening.

Exogenous sucrose influences KEA1 and KEA2 to regulate abscisic acid-mediated primary root growth in Arabidopsis.

Arabidopsis K+-efflux antiporter (KEA)1 and KEA2 are chloroplast inner envelope membrane K+/H+ antiporters that play an important role in plastid development and seedling growth. However, the function of KEA1 and KEA2 during early seedling development is poorly understood. In this work, we found that in Arabidopsis, KEA1 and KEA2 mediated primary root growth by regulating photosynthesis and the ABA signaling pathway. Phenotypic analyses revealed that in the absence of sucrose, the primary root length of the kea1kea2 mutant was significantly shorter than that of the wild-type Columbia-0 (Col-0) plant. However, this phenotype could be remedied by the external application of sucrose. Meanwhile, HPLC-MS/MS results showed that in sucrose-free medium, ABA accumulation in the kea1kea2 mutant was considerably lower than that in Col-0. Transcriptome analysis revealed that many key genes involved in ABA signals were repressed in the kea1kea2 mutant. We concluded that KEA1 and KEA2 deficiency not only affected photosynthesis but was also involved in primary root growth likely through an ABA-dependent manner. This study confirmed the new function of KEA1 and KEA2 in affecting primary root growth.

Loss of inner-envelope K+/H+ exchangers impairs plastid rRNA maturation and gene expression.

The inner-envelope K+ EFFLUX ANTIPORTERS (KEA) 1 and 2 are critical for chloroplast development, ion homeostasis, and photosynthesis. However, the mechanisms by which changes in ion flux across the envelope affect organelle biogenesis remained elusive. Chloroplast development requires intricate coordination between the nuclear genome and the plastome. Many mutants compromised in plastid gene expression (PGE) display a virescent phenotype, that is delayed greening. The phenotypic appearance of Arabidopsis thaliana kea1 kea2 double mutants fulfills this criterion, yet a link to PGE has not been explored. Here, we show that a simultaneous loss of KEA1 and KEA2 results in maturation defects of the plastid ribosomal RNAs. This may be caused by secondary structure changes of rRNA transcripts and concomitant reduced binding of RNA-processing proteins, which we documented in the presence of skewed ion homeostasis in kea1 kea2. Consequently, protein synthesis and steady-state levels of plastome-encoded proteins remain low in mutants. Disturbance in PGE and other signs of plastid malfunction activate GENOMES UNCOUPLED 1-dependent retrograde signaling in kea1 kea2, resulting in a dramatic downregulation of GOLDEN2-LIKE transcription factors to halt expression of photosynthesis-associated nuclear-encoded genes (PhANGs). PhANG suppression delays the development of fully photosynthesizing kea1 kea2 chloroplasts, probably to avoid progressing photo-oxidative damage. Overall, our results reveal that KEA1/KEA2 function impacts plastid development via effects on RNA-metabolism and PGE.

Mitochondrial K+ Transport: Modulation and Functional Consequences.

The existence of a K+ cycle in mitochondria has been predicted since the development of the chemiosmotic theory and has been shown to be crucial for several cellular phenomena, including regulation of mitochondrial volume and redox state. One of the pathways known to participate in K+ cycling is the ATP-sensitive K+ channel, MitoKATP. This channel was vastly studied for promoting protection against ischemia reperfusion when pharmacologically activated, although its molecular identity remained unknown for decades. The recent molecular characterization of MitoKATP has opened new possibilities for modulation of this channel as a mechanism to control cellular processes. Here, we discuss different strategies to control MitoKATP activity and consider how these could be used as tools to regulate metabolism and cellular events.

c-di-AMP, a likely master regulator of bacterial K+ homeostasis machinery, activates a K+ exporter.

bis-(3',5')-cyclic diadenosine monophosphate (c-di-AMP) is a second messenger with roles in virulence, cell wall and biofilm formation, and surveillance of DNA integrity in many bacterial species, including pathogens. Strikingly, it has also been proposed to coordinate the activity of the components of K+ homeostasis machinery, inhibiting K+ import, and activating K+ export. However, there is a lack of quantitative evidence supporting the direct functional impact of c-di-AMP on K+ transporters. To gain a detailed understanding of the role of c-di-AMP on the activity of a component of the K+ homeostasis machinery in B. subtilis, we have characterized the impact of c-di-AMP on the functional, biochemical, and physiological properties of KhtTU, a K+/H+ antiporter composed of the membrane protein KhtU and the cytosolic protein KhtT. We have confirmed c-di-AMP binding to KhtT and determined the crystal structure of this complex. We have characterized in vitro the functional properties of KhtTU and KhtU alone and quantified the impact of c-di-AMP and of pH on their activity, demonstrating that c-di-AMP activates KhtTU and that pH increases its sensitivity to this nucleotide. Based on our functional and structural data, we were able to propose a mechanism for the activation of KhtTU by c-di-AMP. In addition, we have analyzed the impact of KhtTU in its native bacterium, providing a physiological context for the regulatory function of c-di-AMP and pH. Overall, we provide unique information that supports the proposal that c-di-AMP is a master regulator of K+ homeostasis machinery.

Bacterial infection reinforces host metabolic flux from arginine to spermine for NLRP3 inflammasome evasion.

Hosts recognize cytosolic microbial infection via the nucleotide-binding domain-like receptor (NLR) protein family, triggering inflammasome complex assembly to provoke pyroptosis or cytokine-related caspase-1-dependent antimicrobial responses. Pathogens have evolved diverse strategies to antagonize inflammasome activation. Here, Edwardsiella piscicida gene-defined transposon library screening for lactate dehydrogenase (LDH) release in nlrc4-/- bone marrow-derived macrophages (BMDMs) demonstrates that genes clustered in the bacterial arginine metabolism pathway participate in NLRP3 inflammasome inhibition. Blocking arginine uptake or putrescine export significantly relieves NLRP3 inflammasome inhibition, indicating that this bacterium rewires its arginine metabolism network during infection. Moreover, intracellular E. piscicida recruits the host arginine importer (mCAT-1) and putrescine exporter (Oct-2) to bacterium-containing vacuoles, accompanied by reduced arginine and accumulated cytosolic spermine. Neutralizing E. piscicida-induced cytosolic spermine enhancement by spermine synthetase or extracellular spermine significantly alters NLRP3 inflammasome activation. Importantly, accumulated cytosolic spermine inhibits K+ efflux-dependent NLRP3 inflammasome activation. These data highlight the mechanism of bacterial gene-mediated arginine metabolism control for NLRP3 inflammasome evasion.

Functional characterization of proton antiport regulation in the thylakoid membrane.

During photosynthesis, energy is transiently stored as an electrochemical proton gradient across the thylakoid membrane. The resulting proton motive force (pmf) is composed of a membrane potential (ΔΨ) and a proton concentration gradient (ΔpH) and powers the synthesis of ATP. Light energy availability for photosynthesis can change very rapidly and frequently in nature. Thylakoid ion transport proteins buffer the effects that light fluctuations have on photosynthesis by adjusting pmf and its composition. Ion channel activities dissipate ΔΨ, thereby reducing charge recombinations within photosystem II. The dissipation of ΔΨ allows for increased accumulation of protons in the thylakoid lumen, generating the signal that activates feedback downregulation of photosynthesis. Proton export from the lumen via the thylakoid K+ exchange antiporter 3 (KEA3), instead, decreases the ΔpH fraction of the pmf and thereby reduces the regulatory feedback signal. Here, we reveal that the Arabidopsis (Arabidopsis thaliana) KEA3 protein homo-dimerizes via its C-terminal domain. This C-terminus has a regulatory function, which responds to light intensity transients. Plants carrying a C-terminus-less KEA3 variant show reduced feed-back downregulation of photosynthesis and suffer from increased photosystem damage under long-term high light stress. However, during photosynthetic induction in high light, KEA3 deregulation leads to an increase in carbon fixation rates. Together, the data reveal a trade-off between long-term photoprotection and a short-term boost in carbon fixation rates, which is under the control of the KEA3 C-terminus.

Insights into the mechanisms of transport and regulation of the arabidopsis high-affinity K+ transporter HAK51.

The high-affinity K+ transporter HAK5 from Arabidopsis (Arabidopsis thaliana) is essential for K+ acquisition and plant growth at low micromolar K+ concentrations. Despite its functional relevance in plant nutrition, information about functional domains of HAK5 is scarce. Its activity is enhanced by phosphorylation via the AtCIPK23/AtCBL1-9 complex. Based on the recently published three-dimensionalstructure of the bacterial ortholog KimA from Bacillus subtilis, we have modeled AtHAK5 and, by a mutational approach, identified residues G67, Y70, G71, D72, D201, and E312 as essential for transporter function. According to the structural model, residues D72, D201, and E312 may bind K+, whereas residues G67, Y70, and G71 may shape the selective filter for K+, which resembles that of K+shaker-like channels. In addition, we show that phosphorylation of residue S35 by AtCIPK23 is required for reaching maximal transport activity. Serial deletions of the AtHAK5 C-terminus disclosed the presence of an autoinhibitory domain located between residues 571 and 633 together with an AtCIPK23-dependent activation domain downstream of position 633. Presumably, autoinhibition of AtHAK5 is counteracted by phosphorylation of S35 by AtCIPK23. Our results provide a molecular model for K+ transport and describe CIPK-CBL-mediated regulation of plant HAK transporters.

Loss of function of the chloroplast membrane K+/H+ antiporters AtKEA1 and AtKEA2 alters the ROS and NO metabolism but promotes drought stress resilience.

Potassium (K+) exerts key physiological functions such as osmoregulation, stomatal movement, membrane transport, protein synthesis and photosynthesis among others. Previously, it was demonstrated in Arabidopsis thaliana that the loss of function of the chloroplast K+Efflux Antiporters KEA1 and KEA2, located in the inner envelope membrane, provokes inefficient photosynthesis. Therefore, the main goal of this study was to evaluate the potential impact of the loss of function of those cation transport systems in the metabolism of reactive oxygen and nitrogen species (ROS and RNS). Using 14-day-old seedlings from Arabidopsis double knock-out kea1kea2 mutants, ROS metabolism and NO content in roots and green cotyledons were studied at the biochemical level. The loss of function of AtKEA1 and AtKEA2 did not cause oxidative stress but it provoked an alteration of the ROS homeostasis affecting some ROS-generating enzymes. These included glycolate oxidase (GOX) and NADPH-dependent superoxide generation activity, enzymatic and non-enzymatic antioxidants and both NADP-isocitrate dehydrogenase and NADP-malic enzyme activities. NO content, analyzed by confocal laser scanning microscopy (CLSM), was negatively affected in both photosynthetic and non-photosynthetic organs in kea1kea2 mutant seedlings. Furthermore, the S-nitrosoglutathione reductase (GSNOR) protein expression and activity were downregulated in kea1kea2 mutants, whereas the tyrosine nitrated protein profile, analyzed by immunoblot, was unaffected but the relative expression of each immunoreactive band changed. Moreover, kea1kea2 mutants showed an increased photorespiratory pathway and stomata closure, thus promoting a higher resilience to drought stress. Data suggest that the chloroplast osmotic balance and integrity maintained by AtKEA1 and AtKEA2 are necessary to keep the balance of ROS/RNS metabolism. Moreover, these data open new questions about how endogenous NO generation might be affected by the K+/H+ transport located in the chloroplasts.

Plastidial transporters KEA1 and KEA2 at the inner envelope membrane adjust stromal pH in the dark.

Photosynthesis and carbon fixation depend critically on the regulation of pH in chloroplast compartments in the daylight and at night. While it is established that an alkaline stroma is required for carbon fixation, it is not known how alkaline stromal pH is formed, maintained or regulated. We tested whether two envelope transporters, AtKEA1 and AtKEA2, directly affected stromal pH in isolated Arabidopsis chloroplasts using the fluorescent probe 2',7'-bis-(2-carboxyethyl)-5-(and-6)-carboxyfluorescein (BCECF). External K+ -induced alkalinization of the stroma was observed in chloroplasts from wild-type (WT) plants but not from kea1kea2 mutants, suggesting that KEA1 and KEA2 mediate K+ uptake/H+ loss to modulate stromal pH. While light-stimulated alkalinization of the stroma was independent of KEA1 and KEA2, the rate of decay to neutral pH in the dark is delayed in kea1kea2 mutants. However, the dark-induced loss of a pH gradient across the thylakoid membrane was similar in WT and mutant chloroplasts. This indicates that proton influx from the cytosol mediated by envelope K+ /H+ antiporters contributes to adjustment of stromal pH upon light to dark transitions.

Collaboration between NDH and KEA3 Allows Maximally Efficient Photosynthesis after a Long Dark Adaptation.

In angiosperms, the NADH dehydrogenase-like (NDH) complex mediates cyclic electron transport around PSI (CET). K+ Efflux Antiporter3 (KEA3) is a putative thylakoid H+/K+ antiporter and allows an increase in membrane potential at the expense of the ∆pH component of the proton motive force. In this study, we discovered that the chlororespiratory reduction2-1 (crr2-1) mutation, which abolished NDH-dependent CET, enhanced the kea3-1 mutant phenotypes in Arabidopsis (Arabidopsis thaliana). The NDH complex pumps protons during CET, further enhancing ∆pH, but its physiological function has not been fully clarified. The observed effect only took place upon exposure to light of 110 µmol photons m-2 s-1 after overnight dark adaptation. We propose two distinct modes of NDH action. In the initial phase, within 1 min after the onset of actinic light, the NDH-dependent CET engages with KEA3 to enhance electron transport efficiency. In the subsequent phase, in which the ∆pH-dependent down-regulation of the electron transport is relaxed, the NDH complex engages with KEA3 to relax the large ∆pH formed during the initial phase. We observed a similar impact of the crr2-1 mutation in the genetic background of the PROTON GRADIENT REGULATION5 overexpression line, in which the size of ∆pH was enhanced. When photosynthesis was induced at 300 µmol photons m-2 s-1, the contribution of KEA3 was negligible in the initial phase and the ∆pH-dependent down-regulation was not relaxed in the second phase. In the crr2-1 kea3-1 double mutant, the induction of CO2 fixation was delayed after overnight dark adaptation.

Close Temporal Relationship between Oscillating Cytosolic K+ and Growth in Root Hairs of Arabidopsis.

Root hair elongation relies on polarized cell expansion at the growing tip. As a major osmotically active ion, potassium is expected to be continuously assimilated to maintain cell turgor during hair tip growth. However, due to the lack of practicable detection methods, the dynamics and physiological role of K+ in hair growth are still unclear. In this report, we apply the small-molecule fluorescent K+ sensor NK3 in Arabidopsis root hairs for the first time. By employing NK3, oscillating cytoplasmic K+ dynamics can be resolved at the tip of growing root hairs, similar to the growth oscillation pattern. Cross-correlation analysis indicates that K+ oscillation leads the growth oscillations by approximately 1.5 s. Artificially increasing cytoplasmic K+ level showed no significant influence on hair growth rate, but led to the formation of swelling structures at the tip, an increase of cytosolic Ca2+ level and microfilament depolymerization, implying the involvement of antagonistic regulatory factors (e.g., Ca2+ signaling) in the causality between cytoplasmic K+ and hair growth. These results suggest that, in each round of oscillating root hair elongation, the oscillatory cell expansion accelerates on the heels of cytosolic K+ increment, and decelerates with the activation of antagonistic regulators, thus forming a negative feedback loop which ensures the normal growth of root hairs.

A single nucleotide substitution in TaHKT1;5-D controls shoot Na+ accumulation in bread wheat.

Improving salinity tolerance in the most widely cultivated cereal, bread wheat (Triticum aestivum L.), is essential to increase grain yields on saline agricultural lands. A Portuguese landrace, Mocho de Espiga Branca accumulates up to sixfold greater leaf and sheath sodium (Na+ ) than two Australian cultivars, Gladius and Scout, under salt stress in hydroponics. Despite high leaf and sheath Na+ concentrations, Mocho de Espiga Branca maintained similar salinity tolerance compared to Gladius and Scout. A naturally occurring single nucleotide substitution was identified in the gene encoding a major Na+ transporter TaHKT1;5-D in Mocho de Espiga Branca, which resulted in a L190P amino acid residue variation. This variant prevents Mocho de Espiga Branca from retrieving Na+ from the root xylem leading to a high shoot Na+ concentration. The identification of the tissue-tolerant Mocho de Espiga Branca will accelerate the development of more elite salt-tolerant bread wheat cultivars.

Arabidopsis K+ transporter HAK5-mediated high-affinity root K+ uptake is regulated by protein kinases CIPK1 and CIPK9.

The high-affinity K+ transporter HAK5 is the major contributor to root K+ uptake from dilute solutions in K+-starved Arabidopsis plants. Its functionality is tightly regulated and its activity is enhanced under K+ starvation by the transcriptional induction of the AtHAK5 gene, and by the activation of the transporter via the AtCBL1-AtCIPK23 complex. In the present study, the 26 members of the Arabidopsis CIPK protein kinase family were screened in yeast for their capacity to activate HAK5-mediated K+ uptake. Among them, AtCIPK1 was the most efficient activator of AtHAK5. In addition, AtCIPK9, previously reported to participate in K+ homeostasis, also activated the transporter. In roots, the genes encoding AtCIPK1 and AtCIPK9 were induced by K+ deprivation and atcipk1 and atcipk9 Arabidopsis KO mutants showed a reduced AtHAK5-mediated Rb+ uptake. Activation of AtHAK5 by AtCIPK1 did not occur under hyperosmotic stress conditions, where AtCIPK1 function has been shown to be required to maintain plant growth. Taken together, our data contribute to the identification of the complex regulatory networks that control the high-affinity K+ transporter AtHAK5 and root K+ uptake.

Solution structure of the cytoplasmic domain of NhaP2 a K+/H+ antiporter from Vibrio cholera.

NhaP2 is a K+/H+ antiporter from Vibrio cholerae which consists of a transmembrane domain and a cytoplasmic domain of approximately 200 amino acids, both of which are required for cholera infectivity. Here we present the solution structure for a 165 amino acid minimal cytoplasmic domain (P2MIN) form of the protein. The structure reveals a compact N-terminal domain which resembles a Regulator of Conductance of K+ channels (RCK) domain connected to a more open C-terminal domain via a flexible 20 amino acid linker. NMR titration experiments showed that the protein binds ATP through its N-terminal domain, which was further supported by waterLOGSY and Saturation Transfer Difference NMR experiments. The two-domain organisation of the protein was confirmed by BIOSAXS, which also revealed that there are no detectable-ATP-induced conformational changes in the protein structure. Finally, in contrast to all known RCK domain structures solved to date, the current work shows that the protein is a monomer.

Engineering salinity tolerance in plants: progress and prospects.

MAIN CONCLUSION: There is a need to integrate conceptual framework based on the current understanding of salt stress responses with different approaches for manipulating and improving salt tolerance in crop plants. Soil salinity exerts significant constraints on global crop production, posing a serious challenge for plant breeders and biotechnologists. The classical transgenic approach for enhancing salinity tolerance in plants revolves by boosting endogenous defence mechanisms, often via a single-gene approach, and usually involves the enhanced synthesis of compatible osmolytes, antioxidants, polyamines, maintenance of hormone homeostasis, modification of transporters and/or regulatory proteins, including transcription factors and alternative splicing events. Occasionally, genetic manipulation of regulatory proteins or phytohormone levels confers salinity tolerance, but all these may cause undesired reduction in plant growth and/or yields. In this review, we present and evaluate novel and cutting-edge approaches for engineering salt tolerance in crop plants. First, we cover recent findings regarding the importance of regulatory proteins and transporters, and how they can be used to enhance salt tolerance in crop plants. We also evaluate the importance of halobiomes as a reservoir of genes that can be used for engineering salt tolerance in glycophytic crops. Additionally, the role of microRNAs as critical post-transcriptional regulators in plant adaptive responses to salt stress is reviewed and their use for engineering salt-tolerant crop plants is critically assessed. The potentials of alternative splicing mechanisms and targeted gene-editing technologies in understanding plant salt stress responses and developing salt-tolerant crop plants are also discussed.

H+ Transport by K+ EXCHANGE ANTIPORTER3 Promotes Photosynthesis and Growth in Chloroplast ATP Synthase Mutants.

The composition of the thylakoid proton motive force (pmf) is regulated by thylakoid ion transport. Passive ion channels in the thylakoid membrane dissipate the membrane potential (Δψ) component to allow for a higher fraction of pmf stored as a proton concentration gradient (ΔpH). K+/H+ antiport across the thylakoid membrane via K+ EXCHANGE ANTIPORTER3 (KEA3) instead reduces the ΔpH fraction of the pmf. Thereby, KEA3 decreases nonphotochemical quenching (NPQ), thus allowing for higher light use efficiency, which is particularly important during transitions from high to low light. Here, we show that in the background of the Arabidopsis (Arabidopsis thaliana) chloroplast (cp)ATP synthase assembly mutant cgl160, with decreased cpATP synthase activity and increased pmf amplitude, KEA3 plays an important role for photosynthesis and plant growth under steady-state conditions. By comparing cgl160 single with cgl160 kea3 double mutants, we demonstrate that in the cgl160 background loss of KEA3 causes a strong growth penalty. This is due to a reduced photosynthetic capacity of cgl160 kea3 mutants, as these plants have a lower lumenal pH than cgl160 mutants, and thus show substantially increased pH-dependent NPQ and decreased electron transport through the cytochrome b 6 f complex. Overexpression of KEA3 in the cgl160 background reduces pH-dependent NPQ and increases photosystem II efficiency. Taken together, our data provide evidence that under conditions where cpATP synthase activity is low, a KEA3-dependent reduction of ΔpH benefits photosynthesis and growth.