The Control of Transpiration. Insights from Arabidopsis
Year: 2007
Type: review
Abstract: Stomatal complexes in the epidermes of aerial plant parts are critical sites for the regulation of gas exchange between the plant and the atmosphere. Stomata consist of microscopic pores, each flanked by a pair of guard cells. Guard cells can increase or decrease the size of the pore via changes in their turgor status, hence regulating both CO2 entry into the leaf and transpiration, or the loss of water from the leaf. This Update focuses on recent progress in our understanding of the regulation of transpiration and drought tolerance that has been garnered through the use of Arabidopsis (Arabidopsis thaliana) as a model experimental system. The coordinated regulation of gas exchange is integral to land plant survival because CO2 must be able to penetrate the leaf to allow photosynthesis, yet water loss (transpiration) must be minimized to prevent desiccation, drought stress, and plant death. Transpiration also provides the driving force for the transport of water and nutrients from the roots to the aerial tissues, and the evaporation of water from the substomatal cavity cools the plant (Lambers et al., 1998). While a number of morphological traits can contribute to the overall level of leaf gas exchange (e.g. the density and distribution of stomata, leaf epidermal structure and internal organization, cuticle thickness), the regulation of stomatal aperture size is unique in that it is a dynamic and reversible process by which water loss and CO2 influx can be rapidly fine tuned in response to a number of environmental and intrinsic signals, such as light, CO2, and the plant stress hormone abscisic acid (ABA; Schroeder et al., 2001). Because guard cells integrate and respond to a plethora of signals, they have become a model cell type in the field of plant cell signaling (Blatt, 2000; Schroeder et al., 2001; Roelfsema and Hedrich, 2005). This Update highlights recent research reports on the guard cell physiology of Arabidopsis that include some quantitative measure of stomatal function. These measures include transpiration, stomatal conductance (stomatal conductance is defined as stomatal transpiration divided by the vapor pressure difference between the leaf and the air, and increases with increasing stomatal aperture), leaf water status, and water-use efficiency/transpiration efficiency (the ratio of photosynthetic assimilation to transpiration). By focusing the article in this manner, we hope to promote the synthesis of ideas and approaches between whole-plant physiologists and molecular biologists/geneticists. The former typically measure stomatal regulation of gas exchange and its impact on whole-plant physiology, and may treat the cellular and molecular biology of guard cells as a “black box” that receives and reacts to inputs. The latter typically use model plant species to investigate cell and molecular regulation of guard cell function, and may employ gene expression, stomatal aperture, or a specific guard cell parameter, such as ion fluxes, as a “readout,” without quantifying alterations in gas exchange and concomitant whole-plant impacts. Our premise is that Arabidopsis is an excellent reference plant in which these complementary approaches can be readily combined, and that such an integrated approach has great potential to yield new insights into the biology of transpiration in C3 angiosperms. Arabidopsis is a powerful biological tool for the identification and characterization of the molecular regulators of transpiration because it has a small, sequenced genome and is easy to transform. These characteristics allow researchers to experimentally modulate the levels of candidate regulatory molecules via techniques such as RNA interference, insertional mutagenesis, or genetic overexpression, and many studies that employ such tools are discussed in the following sections. Additionally, the availability of collections of genetic mutants allows for large-scale screens for potential regulators of transpiration and for functional analyses of candidate regulators. For example, one such screen used infrared thermography to detect differences in leaf temperature, a correlate of transpiration, among a collection of Arabidopsis mutants (Merlot et al., 2002; Wang et al., 2004). The screen identified two novel mutations in stomatal regulation, ost1 and ost2; OST1 has been cloned and identified as encoding an Arabidopsis homolog of an ABA-activated protein kinase first identified in Vicia faba and is discussed further below (Li et al., 2000; Mustilli et al., 2002). Quantitative trait loci (QTL) analysis is an alternative to mutant analysis that harnesses naturally occurring variation within a species to identify putative genes and genomic regions involved in the regulation of quantitative traits such as transpiration (Alonso-Blanco and Koornneef, 2000). QTL mapping involves the generation of a segregating population for a particular trait, often either an F2 population or a population of homozygous recombinant inbred lines. The population is then phenotyped for the traits of interest and genotyped using molecular markers. Statistical techniques are then employed to link specific genotypes to traits, which allows for the mapping of traits to particular chromosomal regions. Arabidopsis is a useful species for QTL analysis because of its small size and rapid life cycle; large mapping populations can be grown in a small space and recombinant inbred lines can be generated relatively quickly compared to other species (Alonso-Blanco and Koornneef, 2000). Additionally, once candidate genes of interest are identified, they can be further characterized using the molecular techniques mentioned above. QTL analysis has led to the identification of a number of QTLs affecting transpiration efficiency in Arabidopsis (Juenger et al., 2005; Masle et al., 2005). It will be interesting to see to what extent these loci are found to encode known regulators of stomatal response, such as those discussed in subsequent sections, versus novel regulatory mechanisms. One example of the latter was provided by Masle and colleagues (Masle et al., 2005). Using QTL analysis, they identified one genetic locus, ERECTA, which encodes a Leu-rich repeat receptor-like kinase, as a genetic regulator of transpiration efficiency (Masle et al., 2005). Complementation of genotypes harboring mutations in ERECTA (including the common Arabidopsis ecotype Landsberg erecta) with the wild-type ERECTA allele results in increased transpiration efficiency and reduced stomatal conductance compared to erecta mutants. When plants are drought stressed, the plant hormone ABA accumulates in the shoot, where it both inhibits stomatal opening and promotes stomatal closure, resulting in reduced water loss from the plant. ABA is a key regulator of plant water status and stomatal function, and ABA and drought responses are the focus of the majority of the studies discussed in this Update. It is important to note that the terms drought stress and drought tolerance are used in this review just as they were reported in the original references. In these references, it is usually the case that a plant is deemed drought tolerant if it survives a restricted watering regime. However, if the effect of, e.g. a genetic manipulation, is to reduce transpiration, then, under identical watering regimes, the mutant plant is actually experiencing less drought stress than the wild-type control plant. Research on the effects of altered levels of ABA on transpiration spans several decades, starting with the discovery of the wilty flacca mutant of tomato (Lycopersicon esculentum), which is deficient in ABA synthesis (Tal, 1966). Recent research on this topic has taken advantage of the molecular genetic tools available in the Arabidopsis model system. Production of xanthoxin from epoxycarotenoids is a key step in ABA synthesis (Nambara and Marion-Poll, 2005). A family of seven 9-cis-epoxycarotenoid dioxygenase (NCED) genes is implicated in this process in Arabidopsis, of which NCED3 is most strongly induced by drought (Iuchi et al., 2001). Iuchi and co-authors demonstrated that overexpression of NCED3 resulted in elevated ABA levels, strong induction of the RD29B ABA reporter gene following drought onset, reduced transpiration under well-watered conditions, and improved drought survival. Antisense and T-DNA knockout lines exhibited the opposite phenotypes. ABA levels in the plant reflect a balance between ABA synthesis and ABA catabolism into inactive forms by conjugation or oxidation. ABA oxidation to 8′-hydroxyl ABA (from which spontaneous isomerization to phaseic acid occurs) is catalyzed by four cytochrome P450 monooxygenases in Arabidopsis: CYP707A1 to 4. Of these, CYP707A3 is most strongly induced by ABA during dehydration and rehydration (Umezawa et al., 2006). In a recent study, Shinozaki and colleagues characterized T-DNA insertional mutants and constitutive overexpressing lines of CYP707A3 (Umezawa et al., 2006). The T-DNA mutants exhibited greater ABA content under all conditions, more rapid expression of “classic” markers of ABA-induced gene expression (such as RD29A and RAB18), reduced transpiration, and improved survival after drought treatment. Conversely, CYP707A3-overexpressing lines exhibited lowered ABA content coupled with higher levels of the ABA metabolites phaesic acid and dihydrophaseic acid; these lines exhibited increased transpiration. Interestingly, transgenic alterations in levels of two RING-finger proteins, the RING-H2 protein XERICO and the R2R3-type MYB transcription factor HOS10, strongly affect NCED3 transcript levels, with correlated effects on ABA levels, drought tolerance, and water loss (Zhu et al., 2005; Ko et al., 2006). Numerous genetic mutants in Arabidopsis with alterations in production, sensing, or response to all the major plant hormones provide a wealth of resources with which to investigate hormonal regulation of transpiration. Tanaka and colleagues have used such tools to investigate hormonal cross talk between ABA, ethylene, cytokinins, and auxins in the regulation of stomatal apertures (Tanaka et al., 2005, 2006). When ethylene levels were increased, either via provision of exogenous ethylene or through use of the ethylene-overproducing mutant eto1-1, ABA-induced stomatal closure in epidermal peels was retarded, and greater rates of fresh weight decrease in excised shoots were observed. The effects seem specific to the ABA response, as no alterations in dark-induced stomatal closure were seen. Treatment of epidermal peels with cytokinins (6-benzyladenine or kinetin) or auxins (naphthaleneacetic acid or indole-3-acetic acid) similarly opposed ABA-induced stomatal closure. Tanaka et al. hypothesize that these hormones act indirectly, through enhancement of ethylene production, since the repressive effects of 6-benzyladenine and naphthaleneacetic acid on ABA-induced stomatal closure were negated by genetic (use of the ein3-1 ethylene-insensitive mutant) or pharmacological (application of 1-methylcyclopropene, a competitive inhibitor of ethylene-receptor binding) abrogation of ethylene signaling. These studies illustrate the interconnectivity of hormone signaling in plant systems, an emerging theme in phytohormone research (Gazzarrini and McCourt, 2003; Ko et al., 2006). Stomatal conductance is altered by the opening and closing of stomata, processes which in turn are mediated via changes in the turgor status of the adjacent guard cells. Changes in guard cell turgor result from water influx or efflux into the cell following changes in cell water potential, which arise from alterations in symplastic ion concentrations. Stomatal opening occurs when K+, Cl−, malate2−, and Suc accumulate inside the cells, resulting in water entry into the guard cells and the outbowing and opening of the stomatal pore. Stomatal closure occurs following K+ and anion efflux, resulting in loss of water from the cell, a reduction in cell turgor, and pore closure (Schroeder et al., 2001). Therefore, the channels and transporters responsible for ion transport across cell membranes are key regulators in the control of stomatal aperture and plant water loss. Signals resulting in changes in stomatal aperture alter the activities of a number of ion channels and transporters. For example, ABA can promote stomatal closure and inhibit stomatal opening in part by stimulating an increase in cytosolic Ca2+ levels via activation of plasma membrane and endomembrane Ca2+-permeable channels (Sanders et al., 2002; Fan et al., 2004; and 2004). The increase in cytosolic Ca2+ is a that anion efflux and plasma membrane which inhibits K+ channels and K+ channels (Schroeder et al., 2001). A of of the cell water efflux and closure of the stomatal pore. The major K+ involved in guard cell closure in Arabidopsis is by the gene et al., analyses of mutants that an important in the regulation of transpiration. T-DNA insertional mutants and mutants reduced and dark-induced stomatal closure in epidermal peels and increased water loss from excised compared to wild-type plants et al., analysis that the mutants under conditions, and in transpiration when plants were in the et al., Because K+ influx is critical for stomatal K+ such as are also candidate transpiration regulators. of an Arabidopsis mutant harboring a in found no altered stomatal or regulation of transpiration, genetic may for K+ channels in guard cells et al., 2001; et al., 2001). a number of genes encoding K+ channels are in guard cells of Arabidopsis, and et al., 2001). the effects of functional among K+ Schroeder and colleagues used a approach to decrease the overall level of functional K+ channels in Arabidopsis et al., 2001). plants overexpressing a mutant of reduced K+ and guard cell K+ content et al., 2001). These mutant lines also reduced stomatal reduced water loss from excised and increased water content in following drought stress compared to control a for K+ channels in the regulation of transpiration. In to in cellular two transporters that are in guard cells, and are also involved in the control of transpiration, as regulators of ion et al., et al., 2004). mutants were to ABA of stomatal closure reduced stomatal opening et al., and leaf reduced transpiration in the mutant compared to concomitant with an increase in water-use and mutants reduced water loss from excised and were less wilty than wild-type plants under drought et al., These that in the the reduction in stomatal opening and decrease in transpiration are more important to whole-plant water status than increase in water loss to reduced ABA of stomatal closure. Interestingly, mutants opposite to those of mutants have stomatal apertures in both the and the and increased water loss from excised mutants ABA of stomatal closure et al., 2004). that mutants have increased transpiration and reduced water-use and than type when drought et al., 2004). no genes encoding anion channels involved in stomatal have been identified, of the family are as However, a guard has been to in stomatal opening and plant drought responses et al., mutants no altered to ABA reduced and stomatal opening when is the anion under these conditions, is the anion available to as a ion for K+ of with altered stomatal opening in the mutant et al., Additionally, when grown in mutants are more drought tolerant and have reduced transpiration compared to type et al., Interestingly, wild-type plants more water from excised when was that availability for apertures et al., these that the of in the can affect stomatal regulation and of transpiration, and this effect is in part mediated by into guard cells via the In Arabidopsis, receptor-like genes have been identified, and is that that the may as channels 2002). One putative plant has been implicated in in ABA ABA and control of transpiration et al., 2004). Antisense lines stomatal reduced transpiration and were more drought than wild-type plants et al., 2004). with these these lines also higher transcript levels of ABA genes and higher levels of ABA, as as reduced expression of and which encode regulators of ABA The by which an in gene expression The of stress into cellular and responses is of in both and et al., 2004; et al., 2005; Wang et al., 2005). the identification of for drought and ABA is a major of research in plant biology 2000). It is to to ABA signaling within the of this for a more of this topic in the of guard cell physiology, are several excellent (Blatt, 2000; Schroeder et al., 2001; 2003; Roelfsema and Hedrich, 2005; and 2005). in this we have to the progress that is by focusing on just one of guard cell ABA and the of molecules with which it is found to is first because it is an important regulator of ABA responses and because it is one of the in guard cells. is a type protein The first mutant to be characterized was the mutant et al., et al., et al., This mutant a strong wilty et al., by stomatal conductance et al., 2000). mutants more T-DNA insertional mutants of were et al., et al., et al., 2006). These mutants ABA in stomatal regulation, and this is strongly when mutants are with the genes (Merlot et al., or et al., 2006). loss of results in ABA is characterized as a regulator of ABA of the signaling et al., and of have been of species is in the mutant et al., 2001). Production of is also in the ost1 mutant et al., OST1 of the that in guard cells et al., However, the of OST1 and in the signaling are inhibit and that that and OST1 may of the other OST1 was to with in a and the mutant of inhibits ABA activation of OST1 et al., that OST1 of It is also important to note that these two are e.g. OST1 activation of be a on and also OST1 In the activation of Ca2+-permeable channels the plasma membrane is also et al., as is of cytosolic Ca2+ et al., of anion which in the large anion efflux to stomatal closure, is in plants because these channels are Ca2+ et al., Because all of these responses are in mutant it is to hypothesize that they may be in mutants. also with the transcription factor et al., 2002). discussed in the overexpression studies that is a regulator of ABA-induced gene expression, and it is by that is a regulator of ABA one that the result of ABA activation of of be to inhibit the of this One of the by ABA in guard cells is et al., which a and acid Interestingly, the to and inhibits its et al., 2004). of in Arabidopsis by increases stomatal conductance and drought tolerance et al., 2001; et al., effects that be with loss of of in the guard cells. a mutant of that is to has also results in of ABA-induced stomatal closure et al., 2006). also has in of ABA of K+ channels and stomatal through a that involves the protein et al., Wang et al., 2001; et al., 2003; and 2004; et al., 2006). the is to be et al., are to the for further The has on and of have been identified in guard cells. A that the guard cell signaling for ABA-induced stomatal closure, the has been (Li et al., 2006). the of and biology approaches will be to and of ABA and the by et al. one such approach (Li et al., 2006). Recent in to rapid cellular signaling gene expression changes also in the regulation of stomatal aperture size and water loss in and implicated in drought and ABA regulation of transpiration in Arabidopsis and discussed in this article In this to or to overexpressing lines. and implicated in drought and ABA regulation of transpiration in Arabidopsis and discussed in this article In this to or to overexpressing lines. of a number of genes is by of the ABA-induced genes in the are regulatory in such as protein protein and transcription 2000). One to identify potential regulators of gene expression and of transpiration is to screen for that to such as found in the of a number of ABA genes and 1998). and are that were identified via a screen as et al., 2002). to transgenic lines overexpressing or exhibited drought tolerance and reduced water loss from excised et al., 2002). Conversely, and mutants are more to drought than type et al., 2004). on reporter gene analysis et al., both and are in leaf tissues, guard cells, that they may stomatal in part through regulation of gene expression in guard cells. with this of the and which encode K+ channels that K+ during stomatal are in lines et al., 2002). the protein known as has been to drought tolerance when et al., 2004). However, in this overexpressing a of a reduction in water loss et al., 2005; et al., 2006). drought tolerance may have been because was increased expression of a number of ABA-induced proteins, which are to these illustrate the that plants employ a of to drought tolerance, some of which alterations in stomatal as regulators of ABA signaling may also a in the regulation of transpiration. One such is a protein that with a and known regulator of ABA responses et al., 2002). plants overexpressing increased water loss from excised and reduced stomatal closure following leaf compared to control plants et al., 2002). A of ABA response is an transcription factor that to the and can be by protein kinase a regulator of ABA signaling et al., 2002; et al., 2005). In lines overexpressing ABA-induced of two genes in their was to be These lines also increased water loss from excised drought tolerance compared to and ABA-induced stomatal closure compared to type et al., to the that regulators of ABA Conversely, RNA lines that reduced levels of ABA et al., 2005). Interestingly, in expression of ABA-induced genes by is by the et al., 2005). In with a which may with et al., 2005). This a for and in ABA regulation of gene expression et al., 2005). one of four is also implicated in ABA regulation of gene expression and 2006). plants of the genes RD29B and and reduced transcript levels of a regulator of ABA with these plants also drought tolerance and reduced water loss from excised and 2006). However, overexpression of also reduced ABA in ABA of and that the of in ABA response may and cell involved in the of also a in the regulation of stomatal harboring mutations in genes encoding two of the and ABA et al., 2001; et al., 2004). mutants are to ABA induction of cytosolic Ca2+ in guard cells and stomatal closure, and less than type following drought stress et al., 2001). In the of exogenous ABA, mutants reduced K+ and anion efflux responses that with the reduced stomatal apertures and stomatal under these conditions, and are with to ABA et al., 2002). mutants similarly drought tolerance and have reduced stomatal conductance compared to type et al., 2004). transcription have been known to in ABA regulation of plant the studies are new on the of transcription in the dynamic regulation of stomatal 2000). In new on of and in ABA responses that we have the with to the by which of gene expression in the control of transpiration. This Update has some of the recent progress that is in understanding the control of transpiration the and molecular levels, using Arabidopsis as a model system. hope that this review will increased among researchers this levels of biological and ABA are two environmental that were discussed in in this guard cells respond to a of environmental and that of et al., 2001; et al., CO2 et al., et al., et al., and et al., 2000; et al., 2002; et al., on transpiration, discussed are important to our of this article has on levels from the molecular to the it is important to note that Arabidopsis is found in 2002; and 2006). Arabidopsis is also to be a tool for and studies of plant populations in respond to water availability and other environmental that impact the control of gas exchange et al., 2003; and that were in this Update. to the many research was to space
Source: PLANT PHYSIOLOGY
Authors: Sarah E. Nilson, Sarah M. Assmann
Institution: Pennsylvania State University
Language: English
Subfield: Plant Science
Domain: Life Sciences
Open Access status: green
OpenAlex ID: W2154364602