A novel semi-dominant mutation in brassinosteroid signaling kinase1 increases stomatal density

Stomata play a pivotal role in balancing CO2 uptake for photosynthesis and water loss via transpiration. Thus, appropriate regulation of stomatal movement and its formation are crucial for plant growth and survival. Red and blue light induce phosphorylation of the C-terminal residue of the plasma membrane (PM) H+-ATPase, threonine, in guard cells, generating the driving force for stomatal opening. While significant progress has been made in understanding the regulatory mechanism of PM H+-ATPase in guard cells, the regulatory components for the phosphorylation of PM H+-ATPase have not been fully elucidated. Recently, we established a new immunohistochemical technique for detecting guard-cell PM H+-ATPase phosphorylation using leaves, which was expected to facilitate investigations with a single leaf. In this study, we applied the technique to genetic screening experiment to explore novel regulators for the phosphorylation of PM H+-ATPase in guard cells, as well as stomatal development. We successfully performed phenotyping using a single leaf. During the experiment, we identified a mutant exhibiting high stomatal density, jozetsu (jzt), named after a Japanese word meaning ‘talkative’. We found that a novel semi-dominant mutation in BRASSINOSTEROID SIGNALING KINASE1 (BSK1) is responsible for the phenotype in jzt mutant. The present results demonstrate that the new immunohistochemical technique has a wide range of applications, and the novel mutation would provide genetic tool to expand our understanding of plant development mediated by brassinosteroid signaling

Functional characterization of a constitutively active kinase variant of Arabidopsis phototropin 1

Phototropins (phots) are plasma membrane-associated serine/threonine kinases that coordinate a range of processes linked to optimizing photosynthetic efficiency in plants. These photoreceptors contain two light-, oxygen- or voltage-sensing (LOV) domains within their N-terminus, with each binding one molecule of flavin mononucleotide (FMN) as a UV/blue light absorbing chromophore. Although phots contain two LOV domains, light-induced activation of the C-terminal kinase domain and subsequent receptor autophosphorylation is controlled primarily by the A′α-LOV2-Jα photosensory module. Mutations that disrupt interactions between the LOV2-core and its flanking helical segments can uncouple this mode of light regulation. Yet, the impact of these mutations on phot function in Arabidopsis has not been explored. Here, we report that histidine substitution of Arg-472 located within the A′α-helix of Arabidopsis phot1 to histidine results in constitutively activates kinas activity in vitro without affecting LOV2 photochemistry. Expression analysis of phot1 R472H in the phot-deficient mutant confirmed that it is autophosphorylated in darkness in vivo, but was unable to initiate phot1 signaling in the absence of light. Instead, we found that the phot1 R472H mutant is poorly functional under low-light conditions, but can restore phototropism, chloroplast accumulation, stomatal opening, and leaf positioning and expansion at higher light intensities. Our findings suggest that Arabidopsis can adapt to the elevated phosphorylation status of the phot1 R472H mutant by in part reducing its stability, whereas the activity the mutant under high-light conditions can be attributed to additional increases in LOV2-mediated photoreceptor autophosphorylation

Inhibition of light-induced stomatal opening by allyl isothiocyanate does not require guard cell cytosolic Ca2+ signaling

The glucosinolate-myrosinase system is a well-known defense system that has been shown to induce stomatal closure in Brassicales. Isothiocyanates are highly reactive hydrolysates of glucosinolates, and an isothiocyanate, allyl isothiocyanate (AITC), induces stomatal closure accompanied by elevation of free cytosolic Ca2+ concentration ([Ca2+](cyt)) in Arabidopsis. It remains unknown whether AITC inhibits light-induced stomatal opening. This study investigated the role of Ca2+ in AITC-induced stomatal closure and inhibition of light-induced stomatal opening. AITC induced stomatal closure and inhibited light-induced stomatal opening in a dose-dependent manner. A Ca2+ channel inhibitor, La3+, a Ca(2+)chelator, EGTA, and an inhibitor of Ca2+ release from internal stores, nicotinamide, inhibited AITC-induced [Ca2+](cyt) elevation and stomatal closure, but did not affect inhibition of light-induced stomatal opening. AITC activated non-selective Ca2+-permeable cation channels and inhibited inward-rectifying K+ (K-in(+)) channels in a Ca2+-independent manner. AITC also inhibited stomatal opening induced by fusicoccin, a plasma membrane H+-ATPase activator, but had no significant effect on fusicoccin-induced phosphorylation of the penultimate threonine of H+-ATPase. Taken together, these results suggest that AITC induces Ca2+ influx and Ca2+ release to elevate [Ca2+](cyt), which is essential for AITC-induced stomatal closure but not for inhibition of K-in(+) channels and light-induced stomatal opening

光電界センサを用いたレーザによる放電誘導機構の解明

It is very important to clarify the mechanism of discharge induced by a laser-produced plasma channel as a basic study on the laser-induced lightning. We measured the temporal evolution of the electron density and the neutral particle density in one of plasma beads in the channel using Thomson and Rayleigh scattering methods, and also performed discharge induction experiments for a short gap. It was found that the induced discharge is greatly influenced by the temporal evolution of the electron density and the neutral particle density in a plasma. In this paper, we measured the electric field along the channel using an optical sensor at the discharge induction to study the leader propagation process in a long gap. The induced discharge in a long gap changes in the order of electron avalanche, streamer, leader, and final sparkover similarly as in a long gap discharge in the atmosphere. To clarify the leader propagation, we measured the change in the electric field due to the leader propagation between the electrodes, and estimated the velocity of the leader propagation. The electric field was measured using an optical sensor based on the Pockels effect

Mechanosensory trichome cells evoke a mechanical stimuli–induced immune response in Arabidopsis thaliana

Perception of pathogen-derived ligands by corresponding host receptors is a pivotal strategy in eukaryotic innate immunity. In plants, this is complemented by circadian anticipation of infection timing, promoting basal resistance even in the absence of pathogen threat. Here, we report that trichomes, hair-like structures on the epidermis, directly sense external mechanical forces, including raindrops, to anticipate pathogen infections in Arabidopsis thaliana. Exposure of leaf surfaces to mechanical stimuli initiates the concentric propagation of intercellular calcium waves away from trichomes to induce defence-related genes. Propagating calcium waves enable effective immunity against pathogenic microbes through the CALMODULIN-BINDING TRANSCRIPTION ACTIVATOR 3 (CAMTA3) and mitogen-activated protein kinases. We propose an early layer of plant immunity in which trichomes function as mechanosensory cells that detect potential risks

A conserved signaling axis integrates conicting environmental drought and heat signals to control stomatal aperture in plants

Plants continuously respond to changing environmental conditions to prevent damage and maintain optimal performance. To regulate gas exchange with the environment and to control abiotic stress relief, plants have pores in their leaf epidermis, called stomata 1. Multiple environmental signals affect the opening and closing of these stomata 2. Heat promotes stomatal opening (to cool down) and drought induces stomatal closing (to prevent water loss). Coinciding stress conditions, however, may evoke conflicting stomatal responses, but the cellular mechanisms to resolve these conflicts are unknown. Here, we demonstrate that the high temperature-associated kinase TARGET OF TEMPERATURE 3 (TOT3) directly controls the activity of plasma membrane H+-ATPases to induce stomatal opening. This TOT3 activity is directly antagonized by OPEN STOMATA 1 (OST1), to prevent water loss during drought stress. This signaling axis harmonizes conflicting heat and drought signals to regulate stomatal aperture. In the context of global climate change, understanding how conflicting stress signals converge on stomatal regulation allows the development of climate change-ready crops

Control of seed dormancy and germination by DOG1-AHG1 PP2C phosphatase complex via binding to heme

Abscisic acid (ABA) regulates abiotic stress and developmental responses including regulation of seed dormancy to prevent seeds from germinating under unfavorable environmental conditions. ABA HYPERSENSITIVE GERMINATION1 (AHG1) encoding a type 2C protein phosphatase (PP2C) is a central negative regulator of ABA response in germination; however, the molecular function and regulation of AHG1 remain elusive. Here we report that AHG1 interacts with DELAY OF GERMINATION1 (DOG1), which is a pivotal positive regulator in seed dormancy. DOG1 acts upstream of AHG1 and impairs the PP2C activity of AHG1 in vitro. Furthermore, DOG1 has the ability to bind heme. Binding of DOG1 to AHG1 and heme are independent processes, but both are essential for DOG1 function in vivo. Our study demonstrates that AHG1 and DOG1 constitute an important regulatory system for seed dormancy and germination by integrating multiple environmental signals, in parallel with the PYL/RCAR ABA receptor-mediated regulatory system

Real-world management of treatment-naïve diabetic macular oedema : 2-year visual outcome focusing on the starting year of intervention from STREAT-DMO study

Background/aims To investigate the yearly change of real-world outcomes for best corrected visual acuity (BCVA) after 2-year clinical intervention for treatment-naïve diabetic macular oedema (DMO). Methods Retrospective analysis of aggregated, longitudinal medical records obtained from 27 retina specialised institutions in Japan from Survey of Treatment for DMO database. A total of 2049 treatment-naïve centre involving DMO eyes of which the initial intervention started between 2010 and 2015, and had been followed for 2 years, were eligible. As interventions, antivascular endothelial growth factor (VEGF) agents, local corticosteroids, macular photocoagulation and vitrectomy were defined. In each eye, baseline and final BCVA, the number of each intervention for 2 years was extracted. Each eye was classified by starting year of interventional treatment. Results Although baseline BCVA did not change by year, 2-year improvement of BCVA had been increased, and reached to +6.5 letters in the latest term. There is little difference among starting year about proportions of eyes which BCVA gained >15 letters, in contrast to those which lost >15 letters were decreased by year. The proportion of eyes receiving anti-VEGF therapy was dramatically increased, while those receiving the other therapies were gradually decreased. The proportion of eyes which maintained socially good vision of BCVA>20/40 has been increased and reached to 59.0% in the latest term. Conclusion For recent years, treatment patterns for DMO have been gradually but certainly changed; as a result, better visual gain, suppression of worsened eyes and better final BCVA have been obtained. Anti-VEGF therapy has become the first-line therapy and its injection frequency has been increasing