No abstract

Guard cells surround stomatal pores in the epidermis of plant leaves and stems. Stomatal pore opening is essential for CO2 influx into leaves for photosynthetic carbon fixation. In exchange, plants lose over 95% of their water via transpiration to the atmosphere. Signal transduction mechanisms in guard cells integrate hormonal stimuli, light signals, water status, CO2, temperature, and other environmental conditions to modulate stomatal apertures for regulation of gas exchange and plant survival under diverse conditions. Stomatal guard cells have become a highly developed model system for characterizing early signal transduction mechanisms in plants and for elucidating how individual signaling mechanisms can interact within a network in a single cell. In this review we focus on recent advances in understanding signal transduction mechanisms in guard cells.

Transgenic plants overexpressing the vacuolar H ؉ -pyrophosphatase are much more resistant to high concentrations of NaCl and to water deprivation than the isogenic wild-type strains. These transgenic plants accumulate more Na ؉ and K ؉ in their leaf tissue than the wild type. Moreover, direct measurements on isolated vacuolar membrane vesicles derived from the AVP1 transgenic plants and from wild type demonstrate that the vesicles from the transgenic plants have enhanced cation uptake. The phenotypes of the AVP1 transgenic plants suggest that increasing the vacuolar proton gradient results in increased solute accumulation and water retention. Presumably, sequestration of cations in the vacuole reduces their toxic effects. Genetically engineered drought-and salt-tolerant plants could provide an avenue to the reclamation of farmlands lost to agriculture because of salinity and a lack of rainfall. S alt-and drought-tolerant plants maintain their turgor at low water potentials by increasing the number of solute molecules in the cell (1-3). The active transport of solutes depends on the proton gradients established by proton pumps. In plants, three distinct proton pumps generate proton electrochemical gradients across cell membranes. The P-type ATPase pumps cytoplasmic H Plant vacuoles constitute 40-90% of the total intracellular volume of a mature plant cell (5) and, in concert with the cytosol, generate the cell turgor responsible for growth and plant rigidity. Cell turgor depends on the activity of vacuolar H ϩ pumps that maintain the H ϩ electrochemical gradient across the vacuolar membrane, which permits the secondary active transport of inorganic ions, organic acids, sugars, and other compounds. The accumulation of these solutes is required to maintain the internal water balance (6).In principle, increased vacuolar solute accumulation could confer salt and drought tolerance. The sequestration of ions such as sodium could increase the osmotic pressure of the plant and at the same time reduce the toxic effects of this cation. Exposure to NaCl has been shown to induce the H ϩ transport activity of vacuolar pumps in both salt-tolerant (7, 8) and salt-sensitive plants (9). In principle, enhanced expression of either of the vacuolar proton pumps should increase the sequestration of ions in the vacuole by increasing the availability of protons. However, overexpression of the plant vacuolar H ϩ -ATPase would be difficult because it consists of many subunits, each of which would have to be overexpressed at the correct level in a single transgenic plant to achieve higher activity of the multisubunit complex (10). By contrast, the vacuolar H ϩ -pyrophosphatase of Arabidopsis is encoded by a single gene, AVP1 (11). AVP1 can generate a H Here we show that transgenic plants expressing higher levels of the vacuolar proton-pumping pyrophosphatase, AVP1, are more resistant to salt and drought than are wild-type plants. These resistance phenotypes are associated with increased internal stores of solutes. Materials and MethodsGe...

Guard cells are located in the epidermis of plant leaves, and in pairs surround stomatal pores. These control both the influx of CO2 as a raw material for photosynthesis and water loss from plants through transpiration to the atmosphere. Guard cells have become a highly developed system for dissecting early signal transduction mechanisms in plants. In response to drought, plants synthesize the hormone abscisic acid, which triggers closing of stomata, thus reducing water loss. Recently, central regulators of guard cell abscisic acid signalling have been discovered. The molecular understanding of the guard cell signal transduction network opens possibilities for engineering stomatal responses to control CO2 intake and plant water loss.

Oscillations in cytosolic calcium concentration ([Ca2+]cyt) are central regulators of signal transduction cascades, although the roles of individual [Ca2+]cyt oscillation parameters in regulating downstream physiological responses remain largely unknown. In plants, guard cells integrate environmental and endogenous signals to regulate the aperture of stomatal pores and [Ca2+]cyt oscillations are a fundamental component of stomatal closure. Here we systematically vary [Ca2+]cyt oscillation parameters in Arabidopsis guard cells using a 'calcium clamp' and show that [Ca2+]cyt controls stomatal closure by two mechanisms. Short-term 'calcium-reactive' closure occurred rapidly when [Ca2+]cyt was elevated, whereas the degree of long-term steady-state closure was 'calcium programmed' by [Ca2+]cyt oscillations within a defined range of frequency, transient number, duration and amplitude. Furthermore, in guard cells of the gca2 mutant, [Ca2+]cyt oscillations induced by abscisic acid and extracellular calcium had increased frequencies and reduced transient duration, and steady-state stomatal closure was abolished. Experimentally imposing [Ca2+]cyt oscillations with parameters that elicited closure in the wild type restored long-term closure in gca2 stomata. These data show that a defined window of guard cell [Ca2+]cyt oscillation parameters programs changes in steady-state stomatal aperture.

Cytosolic calcium oscillations control signaling in animal cells, whereas in plants their importance remains largely unknown. In wild-type Arabidopsis guard cells abscisic acid, oxidative stress, cold, and external calcium elicited cytosolic calcium oscillations of differing amplitudes and frequencies and induced stomatal closure. In guard cells of the V-ATPase mutant det3, external calcium and oxidative stress elicited prolonged calcium increases, which did not oscillate, and stomatal closure was abolished. Conversely, cold and abscisic acid elicited calcium oscillations in det3, and stomatal closure occurred normally. Moreover, in det3 guard cells, experimentally imposing external calcium-induced oscillations rescued stomatal closure. These data provide genetic evidence that stimulus-specific calcium oscillations are necessary for stomatal closure.

No abstract

Abscisic acid (ABA) is a plant hormone involved in the response of plants to reduced water availability. Reduction of guard cell turgor by ABA diminishes the aperture of the stomatal pore and thereby contributes to the ability of the plant to conserve water during periods of drought. Previous work has demonstrated that cytosolic Ca 2؉ is involved in the signal transduction pathway that mediates the reduction in guard cell turgor elicited by ABA. Here we report that ABA uses a Ca 2؉ -mobilization pathway that involves cyclic adenosine 5-diphosphoribose (cADPR). Microinjection of cADPR into guard cells caused reductions in turgor that were preceded by increases in the concentration of free Ca 2؉ in the cytosol. Patch clamp measurements of isolated guard cell vacuoles revealed the presence of a cADPRelicited Ca 2؉ -selective current that was inhibited at cytosolic Ca 2؉ > 600 nM. Furthermore, microinjection of the cADPR antagonist 8-NH 2 -cADPR caused a reduction in the rate of turgor loss in response to ABA in 54% of cells tested, and nicotinamide, an antagonist of cADPR production, elicited a dose-dependent block of ABA-induced stomatal closure. Our data provide definitive evidence for a physiological role for cADPR and illustrate one mechanism of stimulus-specific Ca 2؉ mobilization in higher plants. Taken Abscisic acid (ABA) is a plant hormone that plays major roles in the control of development and in the response to various environmental stresses. During periods of reduced water availability ABA builds up in the leaves and promotes reductions in the aperture of the stomatal pore. This reduction in stomatal aperture is beneficial to the plant as it serves to reduce the extent of transpirational water loss (1). The reductions in pore width are caused by decreases in the turgor of the two guard cells that surround the stomatal pore. ABA brings about reductions in turgor by promoting the efflux of potassium salt from the guard cells (2). Although it has been known for some time that ABA can operate in guard cells via Ca -permeable channel types on both the plasma membrane and endomembranes, the roles played by these channels in specific signal transduction pathways are unclear (6, 7). Assignment of a particular class or classes of Ca 2ϩ ion channel as response elements in a given stimulus-response coupling pathway is very important in the context of understanding how stimulus-specificity is encoded. This is especially true in the case of the Ca 2ϩ signature because the dynamic properties of the Ca 2ϩ release channels that participate in the response are likely to characterize its spatiotemporal relationships (8).In many animal cells, intracellular Ca 2ϩ mobilization during signaling is achieved by activation of inositol 1,4,5-trisphosphate (InsP 3 ) receptors and͞or ryanodine receptors (9). The Ca 2ϩ -mobilizing properties of InsP 3 in guard cells are established (10), and a recent report has shown that ABA can induce rapid phosphoinositide turnover in guard cells (11). Ca 2ϩ release with the hallmark ch...