The ABC transporter AtABCB14 is a malate importer and modulates stomatal response to CO2

Abstract

Carbon dioxide uptake and water vapour release in plants occur through stomata, which are formed by guard cells. These cells respond to light intensity, CO2 and water availability, and plant hormones. The predicted increase in the atmospheric concentration of CO2 is expected to have a profound effect on our ecosystem. However, many aspects of CO2-dependent stomatal movements are still not understood. Here we show that the ABC transporter AtABCB14 modulates stomatal closure on transition to elevated CO2. Stomatal closure induced by high CO2 levels was accelerated in plants lacking AtABCB14. Apoplastic malate has been suggested to be one of the factors mediating the stomatal response to CO2 (Refs 4,5) and indeed, exogenously applied malate induced a similar AtABCB14-dependent response as high CO2 levels. In isolated epidermal strips that contained only guard cells, malate-dependent stomatal closure was faster in plants lacking the AtABCB14 and slower in AtABCB14-overexpressing plants, than in wild-type plants, indicating that AtABCB14 catalyses the transport of malate from the apoplast into guard cells. Indeed, when AtABCB14 was heterologously expressed in Escherichia coli and HeLa cells, increases in malate transport activity were observed. We therefore suggest that AtABCB14 modulates stomatal movement by transporting malate from the apoplast into guard cells, thereby increasing their osmotic pressure. University of Science and Technology, Pohang, Korea; [10][11][12] and had a strongly reduced sensitivity to glibenclamide, ABA, calcium and auxin, which are well known to control stomatal movement. We therefore were interested whether AtABCB14 also exhibits a regulatory function in guard cell physiology.AtABCB14 expression, as visualized by the activity of an AtABCB14 promoter::GUS fusion construct, is not restricted to guard cells of leaves only, but is also found in guard cells of stems, flowers and siliques ( Fig. 1a-f). In leaves, GUS activity was also detected in epidermal and at very low levels in mesophyll cells (Fig. 1c). These promoter::GUS expressions corresponded to the transcript levels detected in mesophyll and guard cell protoplasts (Fig. 1g). Transient expression of an 35S::AtABCB14:GFP construct in Arabidopsis protoplasts revealed that AtABCB14 is targeted to the plasma membrane (Fig. 1h, i). AtABCB14:sGFP expressed under the control of the AtABCB14 native promoter was targeted to the plasma membrane of guard cells (Fig. 1n). Coexpression of AtABCB14 with AtAHA2:RFP, a fusion protein of a plasma membrane localized proton pump with a red fluorescent protein 13 , resulted in a perfect co-localization ( Fig. 1j-l). Fractionation of microsomes on a sucrose density gradient further confirmed that AtABCB14:HA protein was targeted to the plasma membrane: the distribution pattern of the protein crossreacting with the HA antibody corresponded to that of AtPDR8, a plasma membrane protein 14 and differed from the patterns of the ER (Bip) and vacuolar markers (γ-TIP) (Fig. 1m). These results indica...

A molecular pathway for CO2 response in Arabidopsis guard cells

“…Although several proteins have been shown to function in CO 2 -regulated stomatal response, some critical components are still missing 5,6,18 . Because membrane transporters play an important role in modulating stomatal response to external signals 5,6,[14][15][16][38][39][40] , we hypothesized that mutations in many of such transporters may affect stomatal responses to environmental signals including CO 2 concentration. We identified a large number of genes encoding potential solute transporters and collected T-DNA insertion mutants for these genes to compile a mutant library.…”

“…For responses to ABA and CaCl 2 , intact leaf epidermis were incubated for 3 h in stomatal opening buffer (5 mM MES, 10 mM KCl, 50 mM CaCl 2 , pH 5.6), and exposed to ABA (0, 1, 10 and 50 mM) and CaCl 2 (0, 2 and 10 mM). Thereafter stomatal apertures were measured 16 .…”

“…If RHC1 is an upstream component of HT1, how does HT1 exert its negative effect on SLAC1? A recent study showed that OST1 loss-offunction mutants are strongly impaired in CO 2 -induced stomatal closing and in HCO 3 À activation of the S-type anion channels 16 . This finding indicates that OST1, a protein kinase that regulates SLAC1 activity in the ABA pathway, may be a central integrator of CO 2 and ABA signal transduction in the regulation of SLAC1 in guard cells.…”

“…Elevated CO 2 concentration activates anion channels, resulting in depolarization of plasma membrane in guard cells, causing efflux of solutes and stomatal closure [7][8][9][10] . Earlier studies in Arabidopsis identified several mutants with defects in CO 2 -induced stomatal movements [11][12][13][14][15][16][17][18][19] . However, most of these mutants including abi1-1, abi2-1, gca2, ost1 and slac1 were insensitive to both CO 2 and abscisic acid (ABA), representing components shared by both signalling pathways 5,20 .…”

Alteration of the Interconversion of Pyruvate and Malate in the Plastid or Cytosol of Ripening Tomato Fruit Invokes Diverse Consequences on Sugar But Similar Effects on Cellular Organic Acid, Metabolism, and Transitory Starch Accumulation

“…This finding led to follow-up work demonstrating that the level of malate played an important role in tomato fruit ripening influencing starch accumulation, total soluble solid levels at ripening, and postharvest properties (Centeno et al, 2011), when taken alongside the results of many recent studies on the control of stomatal aperture by malate (NunesNesi et al, 2007;Lee et al, 2008;Araújo et al, 2011;Penfield et al, 2012), thus expanding the documented biological roles for this acid beyond those previously documented (Fernie and Martinoia, 2009;Meyer et al, 2010). This finding thus suggests that malate metabolism may exert a key influence on the normal ripening and metabolism of tomato fruit.…”