Stomatal Guard Cells Research Articles

In vitro polyploid induction of orchids using oryzalin

A protocol for in vitro polyploid induction using oryzalin was developed for Dendrobium, Epidendrum, Odontioda, and Phalaenopsis orchids (Orchidaceae). Protocorms and protocorm-like bodies (PLBs) in liquid nutrient media were subjected to oryzalin treatments of 14.4, 28.9, and 57.7 μM (w/v) concentrations for 3 and 6 days. Higher concentrations and longer treatment durations lowered the survival rates of the explants, but increased the number of polyploids produced. Stomatal guard cell lengths as measured with digital image analysis software, helped to identify several polypoids from digital images of leaf imprints. The optimal treatments were: 14.4 μM for 6 days in Dendrobium and Odontioda; 57.7 μM for 6 days in Epidendrum; and, 14.4 μM for 3 days in Phalaenopsis.

ABA signaling in stomatal guard cells: lessons from Commelina and Vicia

Abscisic acid (ABA) signaling mechanisms have been studied in a broad variety of plant species using complementary analyses, taking advantage of different methodologies suitable for each plant species. Early studies on ABA biosynthesis using Solanum lycopersicum mutants suggested an importance of ABA synthesis in stomatal closure. To understand ABA signaling in guard cells, cellular, biochemical and electrophysiological studies in Vicia faba and Commelina communis have been conducted, providing fundamental knowledge that was further reconfirmed by molecular genetic studies of Arabidopsis. In this article, examples of stomatal studies in several plants and prospects in ABA research are discussed.

Immunohistochemical Detection of Blue Light-Induced Phosphorylation of the Plasma Membrane H+-ATPase in Stomatal Guard Cells

Blue light (BL) receptor phototropins activate the plasma membrane H(+)-ATPase in guard cells through phosphorylation of a penultimate threonine and subsequent binding of the 14-3-3 protein to the phosphorylated C-terminus of H⁺-ATPase, mediating stomatal opening. To date, detection of the phosphorylation level of the guard cell H⁺-ATPase has been performed biochemically using guard cell protoplasts (GCPs). However, preparation of GCPs from Arabidopsis for this purpose requires >5,000 rosette leaves and takes >8 h. Here, we show that BL-induced phosphorylation of guard cell H⁺-ATPase is detected in the epidermis from a single Arabidopsis rosette leaf via an immunohistochemical method using a specific antibody against the phosphorylated penultimate threonine of H⁺-ATPase. BL-induced phosphorylation of the H⁺-ATPase was detected immunohistochemically in the wild type, but not in a phot1-5 phot2-1 double mutant. Moreover, we found that physiological concentrations of the phytohormone ABA completely inhibited BL-induced phosphorylation of guard cell H⁺-ATPase in the epidermis, and that inhibition by ABA in the epidermis is more sensitive than in GCPs. These results indicate that this immunohistochemical method is very useful for detecting the phosphorylation status of guard cell H⁺-ATPase. Thus, we applied this technique to ABA-insensitive mutants (abi1-1, abi2-1 and ost1-2) and found that ABA had no effect on BL-induced phosphorylation in these mutants. These results indicate that inhibition of BL-induced phosphorylation of guard cell H⁺-ATPase by ABA is regulated by ABI1, ABI2 and OST1, which are known to be early ABA signaling components for a wide range of ABA responses in plants.

Mg-chelatase H subunit affects ABA signaling in stomatal guard cells, but is not an ABA receptor in Arabidopsis thaliana

Mg-chelatase H subunit (CHLH) is a multifunctional protein involved in chlorophyll synthesis, plastid-to-nucleus retrograde signaling, and ABA perception. However, whether CHLH acts as an actual ABA receptor remains controversial. Here we present evidence that CHLH affects ABA signaling in stomatal guard cells but is not itself an ABA receptor. We screened ethyl methanesulfonate-treated Arabidopsis thaliana plants with a focus on stomatal aperture-dependent water loss in detached leaves and isolated a rapid transpiration in detached leaves 1 (rtl1) mutant that we identified as a novel missense mutant of CHLH. The rtl1 and CHLH RNAi plants showed phenotypes in which stomatal movements were insensitive to ABA, while the rtl1 phenotype showed normal sensitivity to ABA with respect to seed germination and root growth. ABA-binding analyses using 3H-labeled ABA revealed that recombinant CHLH did not bind ABA, but recombinant pyrabactin resistance 1, a reliable ABA receptor used as a control, showed specific binding. Moreover, we found that the rtl1 mutant showed ABA-induced stomatal closure when a high concentration of extracellular Ca2+ was present and that a knockout mutant of Mg-chelatase I subunit (chli1) showed the same ABA-insensitive phenotype as rtl1. These results suggest that the Mg-chelatase complex as a whole affects the ABA-signaling pathway for stomatal movements.Electronic supplementary materialThe online version of this article (doi:10.1007/s10265-011-0426-x) contains supplementary material, which is available to authorized users.

Technology of In Vitro production of dihaploid Brassica plants

The conditions of cultivating isolated anthers and microspores are optimized for the purpose of producing haploid rapeseed, white head cabbage, and cauliflower plants under in vitro conditions and the process of direct embryogenesis is studied. It is established that the presence of 1 mg/l kinetin and 1 mg/l naphthaleneacetic acid hormones in nutrient medium B5 stimulated the process of direct embryogenesis in rapeseed as well as the formation of secondary and tertiary embryoids on hypocotyl and leaf segments of sterile sprouts. A cytological analysis of regenerant plants (count of the number of chloroplasts in stomatal guard cells and chromosomes in root meristem) confirmed the haploid nature of the rapeseed plants. Regenerants are obtained from callus tissue of cabbage microspores.

Distribution of Fucosylated Xyloglucans among the Walls of Different Cell Types in Monocotyledons Determined by Immunofluorescence Microscopy

Xyloglucans in the non-lignified primary cell walls of different species of monocotyledons have diverse structures, with widely varying proportions of oligosaccharide units that contain fucosylated side chains (F side chains). To determine whether fucosylated xyloglucans occur in all non-lignified walls in a range of monocotyledon species, we used immunofluorescence microscopy with the monoclonal antibody CCRC-M1. The epitope of this antibody, α-L-Fucp-(1→2)-β-D-Galp, occurs in F side chains. In most non-commelinid monocotyledons, the epitope was found in all non-lignified walls. A similar distribution was found in the palm Phoenix canariensis, which is a member of the basal commelinid order Arecales. However, in the other commelinid orders Zingiberales, Commelinales, and Poales, the occurrence of the epitope was restricted, sometimes occurring in only the phloem walls, but often also in walls of other cell types including stomatal guard and subsidiary cells and raphide idioblasts. No epitope was found in the walls of the commelinids Tradescantia virginiana (Commelinaceae, Commelinales) and Zea mays (Poaceae, Poales), but it occurred in the phloem walls of two other Poaceae species, Lolium multiflorum and L. perenne. The distribution of the epitope is discussed in relation to xyloglucan structures in the different taxa. However, the functional significance of the restricted distributions is unknown.

Analysis of reactive oxygen species in the guard cell of wheat stoma with confocal microscope

Recently, the laser-scanning confocal microscope has become a routine technique and indispensable tool for cell biological studies. Previous studies indicated that reactive oxygen species (ROS) were generated in tobacco epidermal cells with confocal microscope. In the present studies, the probe 2',7'-dichlorof luorescein diacetate (H₂DCF-DA) was used to research the change of ROS in the guard cell of wheat stoma, and catalase (CAT) was used to demonstrate that ROS had been labeled. The laser-scanning mode of confocal microscope was XYT, and the time interval between two sections was 1.6351 s. Sixty optical sections were acquired with the laser-scanning confocal microscope, and CAT (60,000 U mg⁻¹) was added after four optical sections were scanned. Furthermore, the region of interest (ROI) was circled and the fluorescence intensity of ROS was quantified with Leica Confocal Software. The quantitative data were exported and the trend chart was made with software Excell. The results indicated that ROS were produced intracellularly in stomatal guard cells, and the quantified fluorescence intensity of ROS was declined with CAT added. It is a good method to research the instantaneous change of ROS in plant cells with confocal microscope and fluorescence probe H₂DCF-DA.

Ploidy screening of anthurium ( Anthurium andreanum Linden ex André) regenerants derived from anther culture

Anther culture was successfully developed in Anthurium andreanum Linden ex André cv. ‘Tropical’, but resulted in variation in morphology and growth response of regenerants. Ploidy levels varied in different morphological variants. Finding a convenient, rapid, reliable, practical and indirect method to screen and determine anthurium ploidy level is important not only for anthurium, but for ornamentals in general. Regenerants derived from anther culture showing three different ploidy levels (haploids, diploids, triploids) were used in this study. Five indirect methods were used to assess chromosome number: chloroplast number in stomatal guard cells (M1), stomatal length and width ratio (M2), stomatal density (M3), ratio of length and width of leaves (M4), and microspore number per anther (M5). These were compared to chromosome counting as the direct and control method (M6). Through simple regression correlation analysis, when compared to M6, M1 was the most convenient and reliable indirect method to determine the ploidy level. This method was highly and significantly positively correlated to anthurium ploidy level ( r = 0.945; p < 0.01). This method could also be applied much faster than the conventional chromosome method.

AtTPS1-mediated trehalose 6-phosphate synthesis is essential for embryogenic and vegetative growth and responsiveness to ABA in germinating seeds and stomatal guard cells

Trehalose and associated metabolites are part of the sugar signalling system in plants and have profound effects on development. Disruption of the TREHALOSE 6-PHOSPHATE SYNTHASE (TPS1) gene in Arabidopsis results in delayed embryo growth, altered cell wall morphology and carbon metabolism and abortion at the torpedo stage. Here we investigate the role of the TPS1 gene in post-embryonic development using two approaches. In the first we use the seed-specific ABI3 promoter to drive the TPS1 cDNA during embryo development, resulting in rescue of the embryo-lethal tps1 phenotype. Lack of expression from the ABI3::TPS1 transgene in post-germinative tps1 seedlings results in severe growth arrest, accumulation of soluble sugars and starch and leads to an increase in expression of genes related to ABA signalling. In the second approach we use TILLING (targeted induced local lesions in genomes) to generate three weaker, non-embryo-lethal, alleles (tps1-11, tps1-12 and tps1-13) and use these to demonstrate that the TPS1 protein plays a key role in modulating trehalose 6-phosphate (T6P) levels in vegetative tissues of Arabidopsis. All three weaker alleles give a consistent phenotype of slow growth and delayed flowering. Germination of tps1-11, tps1-12 and tps1-13 is hypersensitive to ABA with the degree of hypersensitivity correlating with the decrease in T6P levels in the different alleles. Stomatal pore aperture is regulated by ABA, and this was found to be affected in tps1-12. Our results show that the TPS1 gene product plays an essential role in regulating the growth of vegetative as well as embryogenic tissue in a mechanism involving ABA and sugar metabolism.

Arabidopsis calcium-dependent protein kinase CPK10 functions in abscisic acid- and Ca2+-mediated stomatal regulation in response to drought stress.

Plant calcium-dependent protein kinases (CDPKs) may function as calcium sensors and play important roles in the regulation of plant growth and development and in plant responses to biotic and abiotic stresses. The Arabidopsis (Arabidopsis thaliana) genome encodes 34 CDPKs, and most of them have not been functionally characterized. Here, we report the functional characterization of CPK10 in Arabidopsis response to drought stress. The cpk10 mutant, a T-DNA insertion mutant for the Arabidopsis CPK10 gene, showed a much more sensitive phenotype to drought stress compared with wild-type plants, while the CPK10 overexpression lines displayed enhanced tolerance to drought stress. Induction of stomatal closure and inhibition of stomatal opening by abscisic acid (ABA) and Ca(2+) were impaired in the cpk10 mutants. Using yeast two-hybrid methods, a heat shock protein, HSP1, was identified as a CPK10-interacting protein. The interaction between CPK10 and HSP1 was further confirmed by pull-down and bimolecular fluorescence complementation assays. The HSP1 knockout mutant (hsp1) plants showed a similar sensitive phenotype under drought stress as the cpk10 mutant plants and were similarly less sensitive to ABA and Ca(2+) in regulation of stomatal movements. Electrophysiological experiments showed that ABA and Ca(2+) inhibition of the inward K(+) currents in stomatal guard cells were impaired in the cpk10 and hsp1 mutants. All presented data demonstrate that CPK10, possibly by interacting with HSP1, plays important roles in ABA- and Ca(2+)-mediated regulation of stomatal movements.

The role of plastidial glucose‐6‐phosphate/phosphate translocators in vegetative tissues of Arabidopsis thaliana mutants impaired in starch biosynthesis

Arabidopsis thaliana mutants impaired in starch biosynthesis due to defects in either ADP glucose pyrophosphorylase (adg1-1), plastidic phosphoglucose mutase (pgm) or a new allele of plastidic phosphoglucose isomerase (pgi1-2) exhibit substantial activity of glucose-6-phosphate (Glc6P) transport in leaves that is mediated by a Glc6P/phosphate translocator (GPT) of the inner plastid envelope membrane. In contrast to the wild type, GPT2, one of two functional GPT genes of A. thaliana, is strongly induced in these mutants during the light period. The proposed function of the GPT in plastids of non-green tissues is the provision of Glc6P for starch biosynthesis and/or the oxidative pentose phosphate pathway. The function of GPT in photosynthetic tissues, however, remains obscure. The adg1-1 and pgi1-2 mutants were crossed with the gpt2-1 mutant defective in GPT2. Whereas adg1-1/gpt2-1 was starch-free, residual starch could be detected in pgi1-2/gpt2-1 and was confined to stomatal guard cells, bundle sheath cells and root tips, which parallels the reported spatial expression profile of AtGPT1. Glucose content in the cytosolic heteroglycan increased substantially in adg1-1 but decreased in pgi1-2, suggesting that the plastidic Glc6P pool contributes to its biosynthesis. The abundance of GPT2 mRNA correlates with increased levels of soluble sugars, in particular of glucose in leaves, suggesting induction by the sugar-sensing pathway. The possible function of GPT2 in starch-free mutants is discussed in the background of carbon requirement in leaves during the light-dark cycle.

Nicotinate/nicotinamide mononucleotide adenyltransferase-mediated regulation of NAD biosynthesis protects guard cells from reactive oxygen species in ABA-mediated stomatal movement in Arabidopsis

Nicotinamide adenine dinucleotide (NAD) and its derivative nicotinamide adenine dinucleotide phosphate (NADP) are indispensable co-factors in broad-spectrum metabolic events for the maintenance of cellular homeostasis in all living organisms. In this study, the cellular expression levels of NAD biosynthesis genes in Arabidopsis were investigated. A very high expression of nicotinate/nicotinamide mononucleotide adenyltransferase (NMNAT) was observed in the differentiated stomatal guard cells of the leaf surface. Transcriptional analysis confirmed that several genes in the biosynthesis pathway were also highly expressed in stomatal guard cells. In fact, NAD and NADP metabolisms were investigated during stomatal movement. Importantly, the generation of phytohormone ABA-induced reactive oxygen species, which acts as a signal for stomatal closure, was accompanied by markedly decreased levels of NAD. The ABA-induced oxidative stress caused stomatal cell death in the nmnat mutant. Furthermore, stomata partially lost their ability to close leading to drought susceptibility. The stomata were less responsive to opening cues as well. These results indicate that NAD biosynthesis is involved in protecting guard cells from ABA-induced local oxidative stress via the regulation of NMNAT activity. In this study, it is demonstrated that NMNAT is essential for the maintenance of NAD homeostasis enabling sustainable stomatal movement.

Estimating polyploidy levels in fossil &lt;em&gt;Salix&lt;/em&gt;: A critical review of cell size proxy methods

Studies have used cell size as a proxy to estimate polyploidy levels, the number of chromosome sets in somatic cells, in modern and fossil plant species. This paper critically evaluates these methods by reviewing cell size- and polyploidy-related literature, and provides new cell size data from herbarium material and fossil remains of willow genus Salix. The 40 extant taxa used in the study include most of the polyploidy levels encountered in Salix (2n = 38, 76, 114, 152, 190). Diploid and tetraploid species were morphologically similar to the fossil specimens. Specimens from alpine and arctic regions, forms rarely found in the fossil record, were included to extend the range of polyploidy levels. Measurements taken for this study were on the petiole epidermal cells of fossil and herbarium specimens and the stomatal guard cell complexes of herbarium material. A literature review reveals cell size may not relate only to DNA content, but to a plant’s age, nutritional state and the seasonal timing of organ development. Cell size measurements show that cultivated plants grown at elevations more than 800 m below their original place of growth have a significant increase in cell size. Leaf length-to-width ratio, infrageneric classification, and adaptation to dry or humid environments also correlate with cell size. Cell size proxies for estimating polyploidy levels in fossil willows provide only accurate results if morphologically similar modern plant material from natural habitats is used as a reference for comparison. Leaves should be similar in overall shape, base and apex shape, blade length and width, length-to-width ratio, petiole length, petiole-to-blade length ratio, venation pattern, and margin dentition.

An NAC transcription factor orchestrates multiple features of cell wall development in Medicago truncatula

To identify genes controlling secondary cell wall biosynthesis in the model legume Medicago truncatula, we screened a Tnt1 retrotransposon insertion mutant population for plants with altered patterns of lignin autofluorescence. From more than 9000 R1 plants screened, four independent lines were identified with a total lack of lignin in the interfascicular region. The mutants also showed loss of lignin in phloem fibers, reduced lignin in vascular elements, failure in anther dehiscence and absence of phenolic autofluorescence in stomatal guard cell walls. Microarray and PCR analyses confirmed that the mutations were caused by the insertion of Tnt1 in a gene annotated as encoding a NAM (no apical meristem)-like protein (here designated Medicago truncatula NAC SECONDARY WALL THICKENING PROMOTING FACTOR 1, MtNST1). MtNST1 is the only family member in Medicago, but has three homologs (AtNST1-AtNST3) in Arabidopsis thaliana, which function in different combinations to control cell wall composition in stems and anthers. Loss of MtNST1 function resulted in reduced lignin content, associated with reduced expression of most lignin biosynthetic genes, and a smaller reduction in cell wall polysaccharide content, associated with reduced expression of putative cellulose and hemicellulose biosynthetic genes. Acid pre-treatment and cellulase digestion released significantly more sugars from cell walls of nst1 mutants compared with the wild type. We discuss the implications of these findings for the development of alfalfa (Medicago sativa) as a dedicated bioenergy crop.

Stomatal action directly feeds back on leaf turgor: new insights into the regulation of the plant water status from non-invasive pressure probe measurements

Uptake of CO(2) by the leaf is associated with loss of water. Control of stomatal aperture by volume changes of guard cell pairs optimizes the efficiency of water use. Under water stress, the protein kinase OPEN STOMATA 1 (OST1) activates the guard-cell anion release channel SLOW ANION CHANNEL-ASSOCIATED 1 (SLAC1), and thereby triggers stomatal closure. Plants with mutated OST1 and SLAC1 are defective in guard-cell turgor regulation. To study the effect of stomatal movement on leaf turgor using intact leaves of Arabidopsis, we used a new pressure probe to monitor transpiration and turgor pressure simultaneously and non-invasively. This probe permits routine easy access to parameters related to water status and stomatal conductance under physiological conditions using the model plant Arabidopsis thaliana. Long-term leaf turgor pressure recordings over several weeks showed a drop in turgor during the day and recovery at night. Thus pressure changes directly correlated with the degree of plant transpiration. Leaf turgor of wild-type plants responded to CO(2), light, humidity, ozone and abscisic acid (ABA) in a guard cell-specific manner. Pressure probe measurements of mutants lacking OST1 and SLAC1 function indicated impairment in stomatal responses to light and humidity. In contrast to wild-type plants, leaves from well-watered ost1 plants exposed to a dry atmosphere wilted after light-induced stomatal opening. Experiments with open stomata mutants indicated that the hydraulic conductance of leaf stomata is higher than that of the root-shoot continuum. Thus leaf turgor appears to rely to a large extent on the anion channel activity of autonomously regulated stomatal guard cells.

HvALMT1 from barley is involved in the transport of organic anions

Members of the ALMT gene family contribute to the Al3+ resistance of several plant species by facilitating malate efflux from root cells. The first member of this family to be cloned and characterized, TaALMT1, is responsible for most of the natural variation of Al3+ resistance in wheat. The current study describes the isolation and characterization of HvALMT1, the barley gene with the greatest sequence similarity to TaALMT1. HvALMT1 is located on chromosome 2H which has not been associated with Al3+ resistance in barley. The relatively low levels of HvALMT1 expression detected in root and shoot tissues were independent of external aluminium or phosphorus supply. Transgenic barley plants transformed with the HvALMT1 promoter fused to the green fluorescent protein (GFP) indicated that expression of HvALMT1 was relatively high in stomatal guard cells and in root tissues containing expanding cells. GFP fused to the C-terminus of the full HvALMT1 protein localized to the plasma membrane and motile vesicles within the cytoplasm. HvALMT1 conferred both inward and outward currents when expressed in Xenopus laevis oocytes that were bathed in a range of anions including malate. Both malate uptake and efflux were confirmed in oocyte assays using [14C]malate as a radiotracer. It is suggested that HvALMT1 functions as an anion channel to facilitate organic anion transport in stomatal function and expanding cells.

Constitutive expression of mustard annexin, AnnBj1 enhances abiotic stress tolerance and fiber quality in cotton under stress

Annexins belong to a multigene family of Ca(2+) dependent, phospholipid and cytoskeleton binding proteins. They have been shown to be upregulated under various stress conditions. We generated transgenic cotton plants expressing mustard annexin (AnnBj1), which showed enhanced tolerance towards different abiotic stress treatments like sodium chloride, mannitol, polyethylene glycol and hydrogen peroxide. The tolerance to these treatments was associated with decreased hydrogen peroxide levels and enhanced total peroxidase activity, enhanced content of osmoprotectants- proline and sucrose in transgenic plants. They showed higher retention of total chlorophyll and reduced TBARS in leaf disc assays with stress treatments, and decreased hydrogen peroxide accumulation in the stomatal guard cells when compared to their wild type counterparts. They also showed significantly enhanced fresh weight, relative water content, dry weight under stress. Treatment with sodium chloride resulted in enhanced expression of genes for Delta-pyrroline-5-carboxylase synthetase in leaves, and sucrose phosphate synthase, sucrose synthase and cellulose synthase A in the leaves and fibers of transgenic plants. The transgenic plants maintained normal seed development, fiber quality and cellulose content under stress.

Long‐term relationships among atmospheric CO2, stomata, and intrinsic water use efficiency in individual trees

Leaf-level responses to increases in atmospheric carbon dioxide (CO(2)) concentrations could have large implications for water and carbon cycles. We investigated whether stomatal density, guard cell length, and intrinsic water use efficiency (iWUE) of 27 individual trees growing at the Arnold Arboretum in Boston, Massachusetts have responded to changing environmental conditions over the last 100 years. We examined leaves from 74 herbarium specimens collected from three genera-Acer (maples), Quercus (oaks), and Carpinus (hornbeams)-from 1893 to 2006. During this period, global average atmospheric CO(2) concentrations increased by approximately 29% (86 ppm), and temperatures in Boston increased by 1.8°C. Stomatal density and guard cell length were negatively correlated in oaks and hornbeams. Although stomatal density declined and guard cell length increased over time, the changes were not dependent on the magnitude of changes in CO(2) concentrations. Intrinsic WUE did not change significantly over time. Our findings suggest that iWUE may not respond to changes in CO(2) concentrations over the lifetimes of individual trees, possibly because of compensating changes in stomatal density and guard cell size. We provide an example of a method that can enable researchers to differentiate between genetic and plastic responses to global change in long-lived trees.

Stem growth habit affects leaf morphology and gas exchange traits in soybean

The stem growth habit, determinate or indeterminate, of soybean, Glycine max, varieties affects various plant morphological and developmental traits. The objective of this study is to identify the effect of stem growth habit in soybean on the stomatal conductance of single leaves in relation to their leaf morphology in order to better understand the ecological and agronomic significance of this plant trait. The stomatal conductance of leaves on the main stem was measured periodically under favourable field conditions to evaluate g(max), defined as the maximum stomatal conductance at full leaf expansion, for four varieties of soybean and their respective determinate or indeterminate near isogenic lines (NILs). Leaf morphological traits including stomatal density, guard cell length and vein density were also measured. The value of g(max) ranged from 0.383 to 0.754 mol H(2)O m(-2) s(-1) across all the genotypes for both years. For the four pairs of varieties, the indeterminate lines exhibited significantly greater g(max), stomatal density, numbers of epidermal cells per unit area and total vein length per unit area than their respective determinate NILs in both years. The guard cell length, leaf mass per area and single leaf size all tended to be greater in the determinate types. The variation of g(max) across genotypes and years was well explained by the product of stomatal density and guard cell length (r = 0.86, P < 0.01). The indeterminate stem growth habit resulted in a greater maximum stomatal conductance for soybean than the determinate habit, and this was attributed to the differences in leaf structure. This raises the further hypothesis that the difference in stem growth habit results in different water use characteristics of soybean plants in the field. Stomatal conductance under favourable conditions can be modified by leaf morphological traits.

An inducible, modular system for spatio-temporal control of gene expression in stomatal guard cells

Stomata, flanked by pairs of guard cells, are small pores on the leaf surfaces of plants and they function to control gas exchange between plants and the atmosphere. Stomata will open when water is available to allow for the uptake of carbon dioxide for photosynthesis. During periods of drought, stomata will close to reduce desiccation stress. As such, optimal functioning of stomata will impact on water use efficiency by plants. The development of an inducible, modular system for robust and targeted gene expression in stomatal guard cells is reported here. It is shown that application of ethanol vapour to activate the gene expression system did not affect the ability of stomata to respond to ABA in bioassays to determine the promotion of stomatal closure and the inhibition of stomatal opening. The system that has been developed allows for robust spatio-temporal control of gene expression in all cells of the stomatal lineage, thereby enabling molecular engineering of stomatal function as well as studies on stomatal development.