Erratum: Attenuation of postprandial blood glucose in humans consuming isomaltodextrin: carbohydrate loading studies.

[This corrects the article DOI: 10.1080/16546628.2017.1325306.].

A tonoplast-localized magnesium transporter is crucial for stomatal opening in Arabidopsis under high Mg2+ conditions.

Plant stomata play an important role in CO2 uptake for photosynthesis and transpiration, but the mechanisms underlying stomatal opening and closing under changing environmental conditions are still not completely understood. Through large-scale genetic screening, we isolated an Arabidopsis mutant (closed stomata2 (cst2)) that is defective in stomatal opening. We cloned the causal gene (MGR1/CST2) and functionally characterized this gene. The mutant phenotype was caused by a mutation in a gene encoding an unknown protein with similarities to the human magnesium (Mg2+ ) efflux transporter ACDP/CNNM. MGR1/CST2 was localized to the tonoplast and showed transport activity for Mg2+ . This protein was constitutively and highly expressed in guard cells. Knockout of this gene resulted in stomatal closing, decreased photosynthesis and growth retardation, especially under high Mg2+ conditions, while overexpression of this gene increased stomatal opening and tolerance to high Mg2+ concentrations. Furthermore, guard cell-specific expression of MGR1/CST2 in the mutant partially restored its stomatal opening. Our results indicate that MGR1/CST2 expression in the leaf guard cells plays an important role in maintaining cytosolic Mg2+ concentrations through sequestering Mg2+ into vacuoles, which is required for stomatal opening, especially under high Mg2+ conditions.

A 265-Nanometer High-Power Deep-UV Light-Emitting Diode Rapidly Inactivates SARS-CoV-2 Aerosols.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection (COVID-19) is an acute respiratory infection transmitted by droplets, aerosols, and contact. Controlling the spread of COVID-19 and developing effective decontamination options are urgent issues for the international community. Here, we report the quantitative inactivation of SARS-CoV-2 in liquid and aerosolized samples by a state-of-the-art, high-power, AlGaN-based, single-chip compact deep-UV (DUV) light-emitting diode (LED) that produces a record continuous-wave output power of 500 mW at its peak emission wavelength of 265 nm. Using this DUV-LED, we observed a greater-than-5-log reduction in infectious SARS-CoV-2 in liquid samples within very short irradiation times (<0.4 s). When we quantified the efficacy of the 265-nm DUV-LED in inactivating SARS-CoV-2, we found that DUV-LED inactivation of aerosolized SARS-CoV-2 was approximately nine times greater than that of SARS-CoV-2 suspension. Our data demonstrate that this newly developed, compact, high-power 265-nm DUV-LED irradiation system with remarkably high inactivation efficiency for aerosolized SARS-CoV-2 could be an effective and practical tool for controlling SARS-CoV-2 spread. IMPORTANCE We developed a 265-nm high-power DUV-LED irradiation system and quantitatively demonstrated that the DUV-LED can inactivate SARS-CoV-2 in suspensions and aerosols within very short irradiation times. We also found that the inactivation effect was about nine times greater against aerosolized SARS-CoV-2 than against SARS-CoV-2 suspensions. The DUV-LED has several advantages over conventional LEDs and mercury lamps, including high power, compactness, and environmental friendliness; its rapid inactivation of aerosolized SARS-CoV-2 opens up new possibilities for the practical application of DUV-LEDs in high-efficiency air purification systems to control airborne transmission of SARS-CoV-2.

Type 2C protein phosphatase clade D family members dephosphorylate guard cell plasma membrane H+-ATPase.

Plasma membrane (PM) H+-ATPase in guard cells is activated by phosphorylation of the penultimate residue, threonine (Thr), in response to blue and red light, promoting stomatal opening. Previous in vitro biochemical investigation suggested that Mg2+- and Mn2+-dependent membrane-localized type 2C protein phosphatase (PP2C)-like activity mediates the dephosphorylation of PM H+-ATPase in guard cells. PP2C clade D (PP2C.D) was later demonstrated to be involved in PM H+-ATPase dephosphorylation during auxin-induced cell expansion in Arabidopsis (Arabidopsis thaliana). However, it is unclear whether PP2C.D phosphatases are involved in PM H+-ATPase dephosphorylation in guard cells. Transient expression experiments using Arabidopsis mesophyll cell protoplasts revealed that all PP2C.D isoforms dephosphorylate the endogenous PM H+-ATPase. We further analyzed PP2C.D6/8/9, which display higher expression levels than other isoforms in guard cells, observing that pp2c.d6, pp2c.d8, and pp2c.d9 single mutants showed similar light-induced stomatal opening and phosphorylation status of PM H+-ATPase in guard cells as Col-0. In contrast, the pp2c.d6/9 double mutant displayed wider stomatal apertures and greater PM H+-ATPase phosphorylation in response to blue light, but delayed dephosphorylation of PM H+-ATPase in guard cells; the pp2c.d6/8/9 triple mutant showed similar phenotypes to those of the pp2c.d6/9 double mutant. Taken together, these results indicate that PP2C.D6 and PP2C.D9 redundantly mediate PM H+-ATPase dephosphorylation in guard cells. Curiously, unlike auxin-induced cell expansion in seedlings, auxin had no effect on the phosphorylation status of PM H+-ATPase in guard cells.

CIPK23 regulates blue light-dependent stomatal opening in Arabidopsis thaliana.

Phototropins (phot1 and phot2) are plant blue light receptor kinases that function to mediate phototropism, chloroplast movement, leaf flattening, and stomatal opening in Arabidopsis. Considerable progress has been made in understanding the mechanisms associated with phototropin receptor activation by light. However, the identities of phototropin signaling components are less well understood by comparison. In this study, we specifically searched for protein kinases that interact with phototropins by using an in vitro screening method (AlphaScreen) to profile interactions against an Arabidopsis protein kinase library. We found that CBL-interacting protein kinase 23 (CIPK23) interacts with both phot1 and phot2. Although these interactions were verified by in vitro pull-down and in vivo bimolecular fluorescence complementation assays, CIPK23 was not phosphorylated by phot1, as least in vitro. Mutants lacking CIPK23 were found to exhibit impaired stomatal opening in response to blue light but no deficits in other phototropin-mediated responses. We further found that blue light activation of inward-rectifying K+ (K+ in ) channels was impaired in the guard cells of cipk23 mutants, whereas activation of the plasma membrane H+ -ATPase was not. The blue light activation of K+ in channels was also impaired in the mutant of BLUS1, which is one of the phototropin substrates in guard cells. We therefore conclude that CIPK23 promotes stomatal opening through activation of K+ in channels most likely in concert with BLUS1, but through a mechanism other than activation of the H+ -ATPase. The role of CIPK23 as a newly identified component of phototropin signaling in stomatal guard cells is discussed.

Subwavelength high-performance polarizers in the deep ultraviolet region.

This study shows that high-performance metasurface polarizers are possible in the deep ultraviolet (DUV) region. A metasurface consisting of a trilayer of an Al (Aluminum) grating, an Al2O3 (Alumina) thin film, and an Al thin film serves as a mirror with high reflectance for light with polarization parallel to the grooves. The reflectance for polarization perpendicular to the grooves vanishes owing to destructive interference between the propagating and gap surface plasmon polaritons. As a result, the metasurface plays the role of a polarizer with a high extinction ratio exceeding 6.2 × 106. This study also shows that the bandwidth is improved by adjusting the incident angle, period, and the Al2O3 layer of the metasurface. The performance of the metasurface polarizer is discussed experimentally. Our finding paves the way for realizing high-performance polarizers in the DUV region.

Phototropin2 Contributes to the Chloroplast Avoidance Response at the Chloroplast-Plasma Membrane Interface.

Blue-light-induced chloroplast movements play an important role in maximizing light utilization for photosynthesis in plants. Under a weak light condition, chloroplasts accumulate to the cell surface to capture light efficiently (chloroplast accumulation response). Conversely, chloroplasts escape from strong light and move to the side wall to reduce photodamage (chloroplast avoidance response). The blue light receptor phototropin (phot) regulates these chloroplast movements and optimizes leaf photosynthesis by controlling other responses in addition to chloroplast movements. Seed plants such as Arabidopsis (Arabidopsis thaliana) have phot1 and phot2. They redundantly mediate phototropism, stomatal opening, leaf flattening, and the chloroplast accumulation response. However, the chloroplast avoidance response is induced by strong blue light and regulated primarily by phot2. Phots are localized mainly on the plasma membrane. However, a substantial amount of phot2 resides on the chloroplast outer envelope. Therefore, differentially localized phot2 might have different functions. To determine the functions of plasma membrane- and chloroplast envelope-localized phot2, we tethered it to these structures with their respective targeting signals. Plasma membrane-localized phot2 regulated phototropism, leaf flattening, stomatal opening, and chloroplast movements. Chloroplast envelope-localized phot2 failed to mediate phototropism, leaf flattening, and the chloroplast accumulation response but partially regulated the chloroplast avoidance response and stomatal opening. Based on the present and previous findings, we propose that phot2 localized at the interface between the plasma membrane and the chloroplasts is required for the chloroplast avoidance response and possibly for stomatal opening as well.

Raf-like kinases CBC1 and CBC2 negatively regulate stomatal opening by negatively regulating plasma membrane H+-ATPase phosphorylation in Arabidopsis.

Stomatal pores, which are surrounded by pairs of guard cells in the plant epidermis, regulate gas exchange between plants and the atmosphere, thereby controlling photosynthesis and transpiration. Blue light works as a signal to guard cells, to induce intracellular signaling and open stomata. Blue light receptor phototropins (phots) are activated by blue light; phot-mediated signals promote plasma membrane (PM) H+-ATPase activity via C-terminal Thr phosphorylation, serving as the driving force for stomatal opening in guard cells. However, the details of this signaling process are not fully understood. In this study, through an in vitro screening of phot-interacting protein kinases, we obtained the CBC1 and CBC2 that had been reported as signal transducers in stomatal opening. Promoter activities of CBC1 and CBC2 indicated that both genes were expressed in guard cells. Single and double knockout mutants of CBC1 and CBC2 showed no lesions in the context of phot-mediated phototropism, chloroplast movement, or leaf flattening. In contrast, the cbc1cbc2 double mutant showed larger stomatal opening under both dark and blue light conditions. Interestingly, the level of phosphorylation of C-terminal Thr of PM H+-ATPase was higher in double mutant guard cells. The larger stomatal openings of the double mutant were effectively suppressed by the phytohormone abscisic acid (ABA). CBC1 and CBC2 interacted with BLUS1 and PM H+-ATPase in vitro. From these results, we conclude that CBC1 and CBC2 act as negative regulators of stomatal opening, probably via inhibition of PM H+-ATPase activity.

Highly Deep Ultraviolet-Transparent h-BN Film Deposited on an Al0.7Ga0.3N Template by Low-Temperature Radio-Frequency Sputtering.

Hexagonal boron nitride (h-BN) is an attractive wide-bandgap material for application to emitters and detectors operating in the deep ultraviolet (DUV) spectral region. The optical transmittance of h-BN in the DUV region is particularly important for these devices. We report on the deposition of thick h-BN films (>200 nm) on Al0.7Ga0.3N templates via radio-frequency sputtering, along with the realization of ultrahigh transmittance in the DUV region. The fraction of the gas mixture (Ar/N2) was varied to investigate its effects on the optical transmittance of BN. DUV light transmittance of as high as 94% was achieved at 265 nm. This value could be further enhanced to exceed 98% by a post-annealing treatment at 800 °C in a N2 ambient for 20 min. The phase of the highly DUV-transparent BN film was determined to be a purely hexagonal structure via Raman spectra measurements. More importantly, these deposition processes were performed at a low temperature (300 °C), which can provide protection from device performance degradation when applied to actual devices.

Brassinosteroid Induces Phosphorylation of the Plasma Membrane H+-ATPase during Hypocotyl Elongation in Arabidopsis thaliana.

Brassinosteroids (BRs) are steroid phytohormones that regulate plant growth and development, and promote cell elongation at least in part via the acid-growth process. BRs have been suggested to induce cell elongation by the activating plasma membrane (PM) H+-ATPase. However, the mechanism by which BRs activate PM H+-ATPase has not been clarified. In this study, we investigated the effects of BR on hypocotyl elongation and the phosphorylation status of a penultimate residue, threonine, of PM H+-ATPase, which affects the activation, in the etiolated seedlings of Arabidopsis thaliana. Brassinolide (BL), an active endogenous BR, induced hypocotyl elongation, phosphorylation of the penultimate, threonine residue of PM H+-ATPase, and binding of the 14-3-3 protein to PM H+-ATPase in the endogenous BR-depleted seedlings. Changes in both BL-induced elongation and phosphorylation of PM H+-ATPase showed similar concentration dependency. BL did not induce phosphorylation of PM H+-ATPase in the BR receptor mutant bri1-6. In contrast, bikinin, a specific inhibitor of BIN2 that acts as a negative regulator of BR signaling, induced its phosphorylation. Furthermore, BL accumulated the transcripts of SMALL AUXIN UP RNA 9 (SAUR9) and SAUR19, which suppress dephosphorylation of the PM H+-ATPase penultimate residue by inhibiting D-clade type 2C protein phosphatase in the hypocotyls of etiolated seedlings. From these results, we conclude that BL-induced phosphorylation of PM H+-ATPase penultimate residue is mediated via the BRI1-BIN2 signaling pathway, together with the accumulation of SAURs during hypocotyl elongation.

Excess Pyrophosphate within Guard Cells Delays Stomatal Closure.

A variety of cellular metabolic reactions generate inorganic pyrophosphate (PPi) as an ATP hydrolysis byproduct. The vacuolar H+-translocating pyrophosphatase (H+-PPase) loss-of-function fugu5 mutant is susceptible to drought and displays pleotropic postgerminative growth defects due to excess PPi. It was recently reported that stomatal closure after abscisic acid (ABA) treatment is delayed in vhp1-1, a fugu5 allele. In contrast, we found that specific removal of PPi rescued all of the above fugu5 developmental and growth defects. Hence, we speculated that excess PPi itself, rather than vacuolar acidification, might delay stomatal closure. To test this hypothesis, we constructed transgenic plants expressing the yeast IPP1 gene (encoding a cytosolic pyrophosphatase) driven by a guard cell-specific promoter (pGC1::IPP1) in the fugu5 background. Our measurements confirmed stomatal closure defects in fugu5, further supporting a role for H+-PPase in stomatal functioning. Importantly, while pGC1::IPP1 transgenics morphologically mimicked fugu5, stomatal closure was restored in response to ABA and darkness. Quantification of water loss revealed that fugu5 stomata were almost completely insensitive to ABA. In addition, growth of pGC1::IPP1 plants was promoted compared to fugu5 throughout their life; however, it did not reach the wild type level. fugu5 also displayed an increased stomatal index, in violation of the one-cell-spacing rule, and phenotypes recovered upon removal of PPi by pAVP1::IPP1 (FUGU5, VHP1 and AVP1 are the same gene encoding H+-PPase), but not in the pGC1::IPP1 line. Taken together, these results clearly support our hypothesis that dysfunction in stomata is triggered by excess PPi within guard cells, probably via perturbed guard cell metabolism.

Stomatal response to blue light in crassulacean acid metabolism plants Kalanchoe pinnata and Kalanchoe daigremontiana.

Blue light (BL) is a fundamental cue for stomatal opening in both C3 and C4 plants. However, it is unknown whether crassulacean acid metabolism (CAM) plants open their stomata in response to BL. We investigated stomatal BL responses in the obligate CAM plants Kalanchoe pinnata and Kalanchoe daigremontiana that characteristically open their stomata at night and close them for part of the day, as contrasted with C3 and C4 plants. Stomata opened in response to weak BL superimposed on background red light in both intact leaves and detached epidermal peels of K. pinnata and K. daigremontiana. BL-dependent stomatal opening was completely inhibited by tautomycin and vanadate, which repress type 1 protein phosphatase and plasma membrane H+-ATPase, respectively. The plasma membrane H+-ATPase activator fusicoccin induced stomatal opening in the dark. Both BL and fusicoccin induced phosphorylation of the guard cell plasma membrane H+-ATPase in K. pinnata. These results indicate that BL-dependent stomatal opening occurs in the obligate CAM plants K. pinnata and K. daigremontiana independently of photosynthetic CO2 assimilation mode.

Insights into Land Plant Evolution Garnered from the Marchantia polymorpha Genome.

The evolution of land flora transformed the terrestrial environment. Land plants evolved from an ancestral charophycean alga from which they inherited developmental, biochemical, and cell biological attributes. Additional biochemical and physiological adaptations to land, and a life cycle with an alternation between multicellular haploid and diploid generations that facilitated efficient dispersal of desiccation tolerant spores, evolved in the ancestral land plant. We analyzed the genome of the liverwort Marchantia polymorpha, a member of a basal land plant lineage. Relative to charophycean algae, land plant genomes are characterized by genes encoding novel biochemical pathways, new phytohormone signaling pathways (notably auxin), expanded repertoires of signaling pathways, and increased diversity in some transcription factor families. Compared with other sequenced land plants, M. polymorpha exhibits low genetic redundancy in most regulatory pathways, with this portion of its genome resembling that predicted for the ancestral land plant. PAPERCLIP.

Electrical determination of current injection and internal quantum efficiencies in AlGaN-based deep-ultraviolet light-emitting diodes.

We propose a method to determine the current injection efficiency (CIE) and internal quantum efficiency (IQE) of light-emitting diodes (LEDs) during current injection. The method is based on fourth-order polynomial fitting of a modified rate equation to electroluminescence data. Our method can extract the CIE at low injection current densities, unlike conventional methods that generally assume the CIE to be unity. We apply the method to AlGaN-based deep-ultraviolet LEDs. Results show that the CIE was only approximately 51% at low injection current densities and was almost independent of injection current density up to 100 A/cm2. The peak IQE was 77%.

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 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. However, 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 constitutively activates phot1 kinase 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 unable to initiate phot1 signaling in the absence of light. Instead, we found that phot1 R472H 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 in part by reducing its stability, whereas the activity of the mutant under high-light conditions can be attributed to additional increases in LOV2-mediated photoreceptor autophosphorylation.

Attenuation of postprandial blood glucose in humans consuming isomaltodextrin: carbohydrate loading studies.

Background: Isomaltodextrin (IMD) is a novel highly branched α-glucan and its function as a soluble dietary fiber is expected. Objective: The goal of this study was to evaluate the effects of IMD on postprandial glucose excursions in healthy people and to make the mechanism clear. Design: Twenty-nine subjects ingested a solution containing maltodextrin (MD) or sucrose with or without IMD. Fourteen subjects ingested a solution containing glucose with or without IMD. Blood glucose concentrations were then compared between the groups. Furthermore, in vitro digestion, inhibition of digestive enzymes, and glucose absorption tests were conducted. Results: IMD attenuated blood glucose elevation in the subjects with blood glucose excursions at the high end of normal following the ingestion of MD or sucrose or glucose alone. This effect of 5 g IMD was most clear. IMD was digested partially only by small intestinal mucosal enzymes, and maltase and isomaltase activities were weakly inhibited. Furthermore, IMD inhibited the transport of glucose from mucosal side to serosal side. Conclusions: IMD attenuated postprandial blood glucose, after the ingestion of MD or sucrose or glucose. As one of the mechanism, it was suggested that IMD inhibited the absorption of glucose on small intestinal mucosal membrane.

Brassinosteroid Involvement in Arabidopsis thaliana Stomatal Opening.

Stomata within the plant epidermis regulate CO2 uptake for photosynthesis and water loss through transpiration. Stomatal opening in Arabidopsis thaliana is determined by various factors, including blue light as a signal and multiple phytohormones. Plasma membrane transporters, including H+-ATPase, K+ channels and anion channels in guard cells, mediate these processes, and the activities and expression levels of these components determine stomatal aperture. However, the regulatory mechanisms involved in these processes are not fully understood. In this study, we used infrared thermography to isolate a mutant defective in stomatal opening in response to light. The causative mutation was identified as an allele of the brassinosteroid (BR) biosynthetic mutant dwarf5. Guard cells from this mutant exhibited normal H+-ATPase activity in response to blue light, but showed reduced K+ accumulation and inward-rectifying K+ (K+in) channel activity as a consequence of decreased expression of major K+in channel genes. Consistent with these results, another BR biosynthetic mutant, det2-1, and a BR receptor mutant, bri1-6, exhibited reduced blue light-dependent stomatal opening. Furthermore, application of BR to the hydroponic culture medium completely restored stomatal opening in dwarf5 and det2-1 but not in bri1-6. However, application of BR to the epidermis of dwarf5 did not restore stomatal response. From these results, we conclude that endogenous BR acts in a long-term manner and is required in guard cells with the ability to open stomata in response to light, probably through regulation of K+in channel activity.

A Raf-like protein kinase BHP mediates blue light-dependent stomatal opening.

Stomata in the plant epidermis open in response to blue light and affect photosynthesis and plant growth by regulating CO2 uptake and transpiration. In stomatal guard cells under blue light, plasma membrane H+-ATPase is phosphorylated and activated via blue light-receptor phototropins and a signaling mediator BLUS1, and H+-ATPase activation drives stomatal opening. However, details of the signaling between phototropins and H+-ATPase remain largely unknown. In this study, through a screening of specific inhibitors for the blue light-dependent H+-ATPase phosphorylation in guard cells, we identified a Raf-like protein kinase, BLUE LIGHT-DEPENDENT H+-ATPASE PHOSPHORYLATION (BHP). Guard cells in the bhp mutant showed impairments of stomatal opening and H+-ATPase phosphorylation in response to blue light. BHP is abundantly expressed in the cytosol of guard cells and interacts with BLUS1 both in vitro and in vivo. Based on these results, BHP is a novel signaling mediator in blue light-dependent stomatal opening, likely downstream of BLUS1.

Auxin Influx Carrier AUX1 Confers Acid Resistance for Arabidopsis Root Elongation Through the Regulation of Plasma Membrane H+-ATPase.

The plant plasma membrane (PM) H+-ATPase regulates pH homeostasis and cell elongation in roots through the formation of an electrochemical H+ gradient across the PM and a decrease in apoplastic pH; however, the detailed signaling for the regulation of PM H+-ATPases remains unclear. Here, we show that an auxin influx carrier, AUXIN RESISTANT1 (AUX1), is required for the maintenance of PM H+-ATPase activity and proper root elongation. We isolated a low pH-hypersensitive 1 (loph1) mutant by a genetic screen of Arabidopsis thaliana on low pH agar plates. The loph1 mutant is a loss-of-function mutant of the AUX1 gene and exhibits a root growth retardation restricted to the low pH condition. The ATP hydrolysis and H+ extrusion activities of the PM H+-ATPase were reduced in loph1 roots. Furthermore, the phosphorylation of the penultimate threonine of the PM H+-ATPase was reduced in loph1 roots under both normal and low pH conditions without reduction of the amount of PM H+-ATPase. Expression of the DR5:GUS reporter gene and auxin-responsive genes suggested that endogenous auxin levels were lower in loph1 roots than in the wild type. The aux1-7 mutant roots also exhibited root growth retardation in the low pH condition like the loph1 roots. These results indicate that AUX1 positively regulates the PM H+-ATPase activity through maintenance of the auxin accumulation in root tips, and this process may serve to maintain root elongation especially under low pH conditions.