Abscisic Acid (ABA)

ABSCISIC ACID (ABA) Dormancy Germination controls many plant processes including stress responses, development and reproduction

Abscisic Acid Development Biotic stress response

Gene expression Stomatal aperture Stress tolerance

Adapted with permission from RIKEN

• ABA biosynthesis, homeostasis and transport • ABA receptors • Downstream signaling • Whole-plant responses • Guard cell responses • Root developmental responses • Vegetative desiccation responses • Seed desiccation and dormancy • Biotic stress responses

ABA levels increase during stress but decrease when stress is relieved

Jan Zeevaart (1930-2009) was a major contributor to our understanding of ABA synthesis and homeostasis.

Image courtesy of Michigan State University-Department of Energy Plant Research Lab; Zeevaart, J.A.D. (1980). Changes in the levels of abscisic acid and its metabolites in excised leaf blades of Xanthium strumarium during and after water stress. Plant Physiol. 66: 672-678.

Zeaxanthin

Zeaxanthin is abundant in green tissues but can be ABA ABA2 limiting for ABA synthesis in roots

Reprinted from Nambara, E., and Marion-Pol, A. (2003) ABA action and interactions in seeds. Trends Plant Sci. 8: 213-217 with permission from Elsevier.

Zeaxanthin

Antheraxanthin

All trans-Violaxanthin

Schwartz, S.H., Qin, X., and Zeevaart, J.A.D. (2003). Elucidation of the indirect pathway of abscisic acid biosynthesis by mutants, genes, and enzymes. Plant Physiol. 131: 1591-1601.

Wild-type tobacco aba2 mutant

Closed stomata Open stomata Leaf water content

Marin, E., Nussaume, L., Quesada, A., Gonneau, M., Sotta, B., Hugueney, P., Frey, A., and Marion-Poll, A. (1996). Molecular identification of zeaxanthin epoxidase of Nicotiana plumbaginifolia, a gene involved in abscisic acid biosynthesis and corresponding to the ABA locus of Arabidopsis thaliana.EMBO J 15: 2331 – 2342.

9-cis-epoxycarotenoids

Han, S.-Y., Kitahata, N., Sekimata, K., Saito, T., Kobayashi, M., Nakashima, K., Yamaguchi-Shinozaki, K., Shinozaki, K., Yoshida, S., and Asami, T. (2004). A novel inhibitor of 9-cis-epoxycarotenoid dioxygenase in abscisic acid biosynthesis in higher plants. Plant Physiol. 135: 1574-1582.

NCED is 9-cis- epoxycarotenoid dioxygenase

xanthoxin

Leaves from vp14 plants lose water more rapidly than wild- type leaves because of reduced levels of ABA

vp14

Wild-type

Tan, B.C., Schwartz, S.H., Zeevaart, J.A.D., and McCarty, D.R. (1997). Genetic control of abscisic acid biosynthesis in maize. Proc. Natl. Acad. Sci. USA 94: 12235-12240.

Thin-layer chromatography showing cleavage VP14/ of 9-cis-epoxycarotenoids by VP14 NCED

Xanthoxin

VP14 9cV or 9cN By product

From Schwartz, S.H., Tan, B.C., Gage, D.A., Zeevaart, J.A.D., and McCarty, D.R. (1997). Specific oxidative cleavage of carotenoids by VP14 of maize. Science 276: 1872-1874 reprinted with permission from AAAS.

Cluster of NCED genes thought to be involved in ABA synthesis. Some are primarily expressed in seeds, and others in vegetative tissues

Schwartz, S.H., Qin, X., and Zeevaart, J.A.D. (2003). Elucidation of the indirect pathway of abscisic acid biosynthesis by mutants, genes, and enzymes. Plant Physiol. 131: 1591-1601.

Increased mRNA levels are detected within 30 minutes of leaf detachment

During seed Days maturation NCED after pollination expression levels increase and then decrease Audran, C., Borel, C., Frey, A., Sotta, B., Meyer, C., Simonneau, T., and Marion-Poll, A. (1998). Expression studies of the zeaxanthin epoxidase gene in Nicotiana plumbaginifolia. Plant Physiol. 118: 1021-1028; Qin, X., and Zeevaart, J.A.D. (1999). The 9-cis-epoxycarotenoid cleavage reaction is the key regulatory step of abscisic acid biosynthesis in water-stressed bean. Proc. Natl. Acad. Sci. USA 96: 15354-15361.

Control vector PvNCED1 Tobacco plants transformed with an inducible Phaseolis vulgaris NCED gene show increased ABA levels, enhanced drought tolerance and increased seed dormancy

Collectively these studies show that NCED expression is highly correlated with ABA levels and a key regulatory event in ABA synthesis

Qin, X., and Zeevaart, J.A.D. (2002). Overexpression of a 9-cis-epoxycarotenoid dioxygenase gene in Nicotiana plumbaginifolia increases abscisic acid and phaseic acid levels and enhances drought tolerance. Plant Physiol. 128: 544-551.

aao3 mutants aba2 are wilty

WT aao3 aba2 mutants germinate under inappropriate conditions

Gonzalez-Guzman, M., et al. (2002). The short-chain alcohol dehydrogenase ABA2 catalyzes the conversion of santhoxin to abscisic aldehyde. Plant Cell 14: 1833-1846. Schwartz, S.H., Qin, X., and Zeevaart, J.A.D. (2003). Elucidation of the indirect pathway of abscisic acid biosynthesis by mutants, genes, and enzymes. Plant Physiol. 131: 1591-1601. Seo, M., et al. (2000). The Arabidopsis aldehyde oxidase 3 (AAO3) gene product catalyzes the final step in abscisic acid biosynthesis in leaves. Proc. Nalt. Acad. Sci. USA 97: 12908-12913.

Rehydration Developmental signals [ABA]

-Glc

ABA Xanthium ABA accumulates eliminated strumarium

Leaf detached Rehydration to initiation by immersion Rehydration drying at time in water ZERO

ABA

Krochko, J.E., Abrams, G.D., Loewen, M.K., Abrams, S.R., and Cutler, A.J. (1998). (+)-Abscisic Acid 8'-Hydroxylase Is a Cytochrome P450 Monooxygenase. Plant Physiol. 118: 849-860. Zeevaart, J.A.D. (1980). Changes in the levels of abscisic acid and its metabolites in excised leaf blades of Xanthium strumarium during and after water stress. Plant Physiol. 66: 672-678. Charles T. Bryson, USDA Agricultural Research Service, Bugwood.org

ABA ABA converted Xanthium accumulates strumarium to phaseic acid Phaseic Acid

Leaf detached Rehydration to initiation by immersion Rehydration drying at time in water ZERO

ABA

CYP707A genes are upregulated upon transfer to high humidity

Okamoto, M., Tanaka, Y., Abrams, S.R., Kamiya, Y., Seki, M., and Nambara, E. (2009). High humidity induces abscisic acid 8'-hydroxylase in stomata and vasculature to regulate local and systemic abscisic acid responses in Arabidopsis. Plant Physiol. 149: 825-834.

Inhibitors of P450 enzymes can slow the degradation of ABA by ABA 8′- hydroxylase

Reprinted from Kitahata, N., Saito, S., Miyazawa, Y., Umezawa, T., Shimada, Y., Min, Y.K., Mizutani, M., Hirai, N., Shinozaki, K., Yoshida, S., and Asami, T. (2005). Chemical regulation of abscisic acid catabolism in plants by cytochrome P450 inhibitors. Bioorg. Med. Chem. 13: 4491-4498 and Todoroki, Y., Kobayashi, K., Yoneyama, H., Hiramatsu, S., Jin, M.-H., Watanabe, B., Mizutani, M., and Hirai, N. (2008). Structure-activity relationship of uniconazole, a potent inhibitor of ABA 8'-hydroxylase, with a focus on hydrophilic functional groups and conformation. Bioorg. Med. Chem. 16: 3141-3152.with permission from Elsevier

P450 inhibitors are often non- specific. Uniconazole inhibits ABA inactivation AND gibberellin (GA) synthesis, causing growth reduction

Abz-F1 is specific for 8′- hydroxylase and confers drought tolerance without growth inhibition ABA catabolism ABA 8'-hydroxylase (CYP707A)

conformationally restricted

UNI Abz-F1

ent-kaurene oxidase (CYP701A)

GA biosynthesis Abz-F1 Reprinted from Todoroki, Y., Kobayashi, K., Shirakura, M., Aoyama, H., Takatori, K., Nimitkeatkai, H., Jin, M.-H., Hiramatsu, S., Ueno, K., Kondo, S., Mizutani, M., and Hirai, N. (2009). Abscinazole-F1, a conformationally restricted analogue of the plant growth retardant uniconazole and an inhibitor of ABA 8'-hydroxylase CYP707A with no growth-retardant effect. Bioorg. Med. Chem. 17: 6620-6630 with permission from Elsevier.

Loss-of-function of one CYP707A gene copy Wild type reduces germination, and loss-of two copies nearly abolishes it Single mutants

Double mutant

Okamoto, M., Kuwahara, A., Seo, M., Kushiro, T., Asami, T., Hirai, N., Kamiya, Y., Koshiba, T., and Nambara, E. (2006). CYP707A1 and CYP707A2, which encode abscisic acid 8'-hydroxylases, are indispensable for proper control of seed dormancy and germination in Arabidopsis. Plant Physiol. 141: 97-107.

ABA

β-glucosidase ABA glucosyltransferase

-Glc

ABA-glucosyl ester

Reprinted by permission from Macmillan Publishers Ltd: Kushiro, T., Okamoto, M., Nakabayashi, K., Yamagishi, K., Kitamura, S., Asami, T., Hirai, N., Koshiba, T., Kamiya, Y., and Nambara, E. (2004). The Arabidopsis cytochrome P450 CYP707A encodes ABA 8′-hydroxylases: key enzymes in ABA catabolism. EMBO J 23: 1647-1656 copyright 2004.

AtBG1

-Glc

Reprinted from Schroeder, J.I., and Nambara, E. (2006). A quick release mechanism for abscisic acid. Cell 126: 1023-1025 with permission from Elsevier.

-Glc

Reprinted from Lee, K.H., Piao, H.L., Kim, H.-Y., Choi, S.M., Jiang, F., Hartung, W., Hwang, I., Kwak, J.M., Lee, I.-J., and Hwang, I. (2006). Activation of gucosidase via stress-induced polymerization rapidly increases active pools of abscisic acid. Cell 126: 1109-1120 with permission from Elsevier.

Plants deficient for UDP glucosyltransferase UGT71C5 have less glucosyl ester and more free ABA. and are more X resistant to drought stress than wild type plants

-Glc

Liu, Z., Yan, J., Li, D., Luo, Q., Yan, Q., Liu, Z., Ye, L., Wang, J., Li, X. and Yang, Y. (2015). UGT71C5, a major glucosyltransferase mediates ABA homeostasis in Arabidopsis thaliana. Plant Physiol. 167: 1659-1670.

NCED 9-cis- xanthoxin Rehydration expoxycarotenoids Developmental signals

Water stress [ABA] Developmental signals

-Glc

ABA translocation and root hydraulic signals may be involved in signaling from root to shoot Well-watered plant with Water-stressed open stomata plant with closed and high stomata and low transpiration transpiration rate rate

Well-watered plant with Exposing part of the root open stomata system to dry conditions and high reduces stomatal aperture transpiration and water use without rate inducing detrimental drought stress effects

A reporter construct made of an ABA-inducible promoter fused to luciferase was used to image ABA levels Luciferase image Luciferase image merged with bright field image

Christmann, A., Hoffmann, T., Teplova, I., Grill, E., and Muller, A. (2005). Generation of active pools of abscisic acid revealed by in vivo imaging of water-stressed Arabidopsis. Plant Physiol. 137: 209-219.

Cell wall pH 5.5

As a weak acid, abscisic acid is a Cytoplasm pH 7 charged anion (ABA-) in the cytoplasm (pH 7) ABA-

In the more acidic cell wall (pH 5.5) some is uncharged (ABAH). This ABAH ABA- + H+ presumably enhances the movement of ABA into but not out of cells ABAH

ABA- + H+

AtABCG25pro::GUS expression

AtABCG25 is Control expressed in veins - ABA and encodes an ABA exporter

10 μM ABA AtABCG25 is transcriptionally induced by ABA

Kuromori, T., Miyaji, T., Yabuuchi, H., Shimizu, H., Sugimoto, E., Kamiya, A., Moriyama, Y., and Shinozaki, K. (2010) ABC transporter AtABCG25 is involved in abscisic acid transport and responses. Proc. Natl. Acad. Sci. USA 107: 2361-2366.

ABA-

Guard cells in loss-of- ABA-induced function abcg40 mutants stomatal closure are less sensitive to ABA, rendering the mutants more susceptible to drought stress

Kang, J., Hwang, J.-U., Lee, M., Kim, Y.-Y., Assmann, S.M., Martinoia, E., and Lee, Y. (2010) PDR-type ABC transporter mediates cellular uptake of the phytohormone abscisic acid. Proc. Natl. Acad. Sci. USA 107: 2355-2360.

EcS = extracellular space ABA importer ABA exporter Cpl = cytoplasm

Vac = vacuole Transporter ABA importer of ABA-GE

ABA exporter ABA importer

Jarzyniak, K.M. and Jasinski, M. (2014). Membrane transporters and drought resistance – a complex issue. Front. Plant Sci. 5: 687.

•In most but not all tissues NCED is rate limiting for ABA synthesis

•ABA synthesis increases with drought stress and during seed maturation

•ABA can be degraded to phaseic acid or reversibly conjugated to ABA-GE

•ABA can be transported within the plant, from root to shoot and from vascular tissues to guard cells

PYR1 PYR/RCAR receptors The core ABA signaling pathway Phosphatase PP2C Protein phosphatases (including ABI1)

Protein kinases (including Kinase SnRK2s and CDPKs)

P P TF

ABA RESPONSES

GFP:RCAR1

PYR/RCAR proteins are found in the cytoplasm and nucleus (arrow) RCAR1pro:GFP

From Ma, Y., Szostkiewicz, I., Korte, A., Moes, D.l., Yang, Y., Christmann, A., and Grill, E. (2009). Regulators of PP2C phosphatase activity function as abscisic acid sensors. Science 324: 1064-1068 reprinted with permission from AAAS.

Wild-type plants fail to germinate on ABA-containing medium Pyrobactin-insensitive mutants are ABA- insensitive and so germinate on ABA- containing medium

The ABA-insensitive mutant abi1 germinates on ABA- containing medium

From Park, S.-Y., et al., and Cutler, S.R. (2009). Abscisic acid inhibits type 2C protein phosphatases via the PYR/PYL family of START proteins. Science 324: 1068-1071 reprinted with permission from AAAS.

© 2015 American Society of Plant Biologists There are many genes encoding PYR/RCARs Number of The 14 PYR/RCARs in Arabidopsis Common Name Species genes Soybean Glycine max 23 Corn Zea mays 20 Populus Western poplar 14 trichocarpa Rice Oryza sativa 11 Grape Vitis vinifera 8 Sorghum Sorghum bicolor 8 Barrel medic (a Medicago 6 model legume) truncatula Arabidopsis Arabidopsis 14 thaliana Klingler, J.P., Batelli, G., and Zhu, J.-K. ABA receptors: the START of a new paradigm in phytohormone signalling. J. Exp.Bot. 61: 3199-3210 by permission of Oxford University Press; Raghavendra, A.S., Gonugunta, V.K., Christmann, A., and Grill, E. (2010) ABA perception and signalling. Trends Plant Sci. 15: 395-401.

PYR1 The genes have different expression PYL1 patterns

But there’s some PYL2 functional redundancy – ABA insensitivity PYL4 increases as more genes are knocked-out PYL5

PYL8

Gonzalez-Guzman, M., Pizzio, G.A., Antoni, R., Vera-Sirera, F., Merilo, E., Bassel, G.W., Fernández, M.A., Holdsworth, M.J., Perez-Amador, M.A., Kollist, H. and Rodriguez, P.L. (2012). Arabidopsis PYR/PYL/RCAR receptors play a major role in quantitative regulation of stomatal aperture and transcriptional response to abscisic acid. Plant Cell 24: 2483–2496.

The ABA receptors are targets of selective proteolysis by ubiquitination. ABA stabilizes the receptors by limiting their polyubiquitination

Low ABA levels enable ABA receptor proteolysis, effectively shutting down ABA responses when ABA levels drop

Irigoyen, M.L., Iniesto, E., Rodriguez, L., Puga, M.I., Yanagawa, Y., Pick, E., Strickland, E., Paz-Ares, J., Wei, N., De Jaeger, G., Rodriguez, P.L., Deng, X.W. and Rubio, V. (2014) Targeted degradation of abscisic acid receptors is mediated by the ubiquitin ligase substrate adaptor DDA1 in Arabidopsis. Plant Cell 26: 712 - 728.

CAR proteins have a Overexpression domain that physically of CAR1 leads ABI1 to enhanced interacts with ABA inhibition of PYL1 PYR/PYL proteins …. shoot growth monomer

PYL1 dimer ..and a Ca- Triple car mutants are less sensitive to dependent growth inhibition by lipid binding ABA C2 domain that anchors them to the plasma membrane Thus, CAR proteins membrane enhance ABA sensitivity

Rodriguez, L., Gonzalez-Guzman, M., Diaz, M., Rodrigues, A., Izquierdo-Garcia, A.C., Peirats-Llobet, M., Fernandez, M.A., Antoni, R., Fernandez, D., Marquez, J.A., Mulet, J.M., Albert, A. and Rodriguez, P.L. (2014) C2-domain abscisic acid-related proteins mediate the interaction of PYR/PYL/RCAR abscisic acid receptors with the plasma membrane and regulate abscisic acid sensitivity in Arabidopsis. Plant Cell 26: 4802–4820.

NO ABA ABA

PYR1 PYR1

PP2C

Reprinted from Raghavendra, A.S., Gonugunta, V.K., Christmann, A., and Grill, E. (2010) ABA perception and signalling. Trends Plant Sci. 15: 395-401 with permission from Elsevier.

FRET = Fluorescence Resonance Energy Transfer

ABA ABA lowers FRET efficiency, which can be measured by light No ABA, energy + ABA, no energy transferred between transferred between emission ratios chromophores chromophores This reduction in the emission of fluorescence is seen here in guard cells as a shift towards blue in the emission ratio scale following treatment with 10 µM ABA over time

Waadt, R., Hitomi, K., Nishimura, N., Hitomi, C., Adams, S.R., Getzoff, E.D. and Schroeder, J.I. (2014) FRET-based reporters for the direct visualization of abscisic acid concentration changes and distribution in Arabidopsis. Elife 3: e01739.

Wild type abi1-1

ABA ABA

Dormancy NO Dormancy (Germination)

NO ABA NO ABA

NO Dormancy NO Dormancy (Germination) (Germination)

From Koornneef, M., Reuling, G., and Karssen, C.M. (1984). The isolation and characterization of abscisic acid-insensitive mutants of Arabidopsis thaliana. Physiologia Plantarum 61: 377-383, with permission from John Wiley and Sons.

ABI1 encodes a PP2C protein phosphatase

Guard cells are not ABA- Wild abi1 responsive type

Leung, J., Bouvier-Durand, M., Morris, P., Guerrier, D., Chefdor, F., and Giraudat, J. (1994). Arabidopsis ABA response gene ABI1: features of a calcium-modulated protein phosphatase. Science 264: 1448-1452; Meyer, K., Leube, M., and Grill, E. (1994). A protein phosphatase 2C involved in ABA signal transduction in Arabidopsis thaliana. Science 264: 1452-1455.

ABI1 abi1-1 ABI1

Kinase Kinase Kinase

P P P P NO ABA ABA NO ABA RESPONSES RESPONSES RESPONSES

-AHG1

Six of the clade A proteins (indicated) have a confirmed role in ABA signaling

Reprinted from Schweighofer, A., Hirt, H., and Meskiene, I. (2004). Plant PP2C phosphatases: emerging functions in stress signaling. Trends Plant Sci. 9: 236-243 with permission from Elsevier. See also Xue, T., Wang, D., Zhang, S., Ehlting, J., Ni, F., Jakab, S., Zheng, C. and Zhong, Y. (2008). Genome-wide and expression analysis of protein phosphatase 2C in rice and Arabidopsis. BMC Genomics. 9: 550.

© 2015 American Society of Plant Biologists PP2Cs interfere with the action of SnRK2 protein kinases NO ABA PYR1 In the absence of ABA, SnRK2 protein kinase activity is ABI1 inhibited by PP2C SnRK2 phosphatases

P NO ABA RESPONSES

This interaction is PYR1 stabilized in the dominant abi1 mutant

abi1-1 ABI1 SnRK2 SnRK2 Yeast two-hybrid assay – when two proteins interact, the yeast cells grow

Umezawa, T., Sugiyama, N., Mizoguchi, M., Hayashi, S., Myouga, F., Yamaguchi-Shinozaki, K., Ishihama, Y., Hirayama, T., and Shinozaki, K. (2009). Type 2C protein phosphatases directly regulate abscisic acid-activated protein kinases in Arabidopsis. Prod. Natl. Acad. Sci. USA 106: 17588-17593.

PYR1

PYR1, ABA and PP2C form a PP2C complex that inactivates PP2C (ABI1)

SnRK2 P This permits SnRK2 activation. SnRK2 Phosphorylation targets include

SnRK2s, ion channels and P P transcription factors TF Ion P channel ABA RESPONSES

© 2015 American Society of Plant Biologists PP2C binds ABA + receptor & SnRK kinase similarly A PP2C tyrosine indicated by interacts with both SnRK2 and PYL+ABA. A serine (S) on both SnRK2 and PYL interacts with PP2S When PYL+ABA binds PP2C, SnRK2 is released and autophosphorylates (S-P_

Reprinted from Soon, F.F., Ng, L.M., Zhou, X.E., West, G.M., Kovach, A., Tan, M.H., Suino-Powell, K.M., He, Y., Xu, Y., Chalmers, M.J., Brunzelle, J.S., Zhang, H., Yang, H., Jiang, H., Li, J., Yong, E.L., Cutler, S., Zhu, J.K., Griffin, P.R., Melcher, K. and Xu, H.E. (2012) Molecular mimicry regulates ABA signaling by SnRK2 kinases and PP2C phosphatases. Science 335: 85–88.

Enhanced binding / sustained switch Environmental, Casein “off” of ABA developmental kinase 2 P responses P P information and (CK2) P signals

Enhanced proteolysis, dampens ABA responses

Vilela, B., Nájar, E., Lumbreras, V., Leung, J. and Pagès, M. (2015) Casein kinase 2 negatively regulates abscisic acid-activated SnRK2s in the core abscisic acid-signaling module. Mol Plant 8: 709–721.

The CPDK-SnRK superfamily of protein kinases

SnRK2

P P TF Ion P channel

The SnRK2 subfamily ABA RESPONSES

Hrabak, E.M., Chan, C.W.M., Gribskov, M., Harper, J.F., Choi, J.H., Halford, N., Kudla, J., Luan, S., Nimmo, H.G., Sussman, M.R., Thomas, M., Walker- Simmons, K., Zhu, J.-K., and Harmon, A.C. (2003). The Arabidopsis CDPK-SnRK superfamily of protein kinases. Plant Physiol. 132: 666-680.

The protein kinase activity of the SnRK2 AAPK is enhanced by ABA

Phosphorylation

PKABA1, an ABA- A dominant inducible SnRK2 negative form of protein kinase from AAPK interferes wheat, accumulates with guard cell in developing seeds response to ABA.

Wheat Vicia faba

Anderberg, R.J., and Walker-Simmons, M.K. (1992). Isolation of a wheat cDNA clone for an abscisic acid-inducible transcript with homology to protein kinases. Proc. Natl. Acad. Sci. USA 89: 10183-10187; Li, J., and Assmann, S.M. (1996). An abscisic acid-activated and calcium-independent protein kinase from guard cells of fava bean. Plant Cell 8: 2359-2368. From Li, J., Wang, X.-Q., Watson, M.B., and Assmann, S.M. (2000). Regulation of abscisic acid-induced stomatal closure and anion channels by guard cell AAPK kinase. Science 287: 300-303 reprinted with permission of AAAS.

Wild type Loss-of-function ost1 mutants have reduced stomatal closure response. OST1 is ost1-1 expressed in guard cells and vascular tissues ost1-2

ost1-1 ost1-2 Wild type

Mustilli, A.-C., Merlot, S., Vavasseur, A., Fenzi, F., and Giraudat, J. (2002). Arabidopsis OST1 protein kinase mediates the regulation of stomatal aperture by abscisic acid and acts upstream of reactive oxygen species production. Plant Cell 14: 3089-3099.

aba2-13

Mutants impaired in ABA synthesis (aba2-13) or signaling (ost1-4) show reduced stomatal closure in response to a decrease in humidity, ost1-4 and so are cooler than wild-type plants

Reprinted from Xie, X., Wang, Y., Williamson, L., Holroyd, G.H., Tagliavia, C., Murchie, E., Theobald, J., Knight, M.R., Davies, W.J., Leyser, H.M.O., and Hetherington, A.M. (2006). The identification of genes involved in the stomatal response to reduced atmospheric relative humidity. Curr Biol. 16: 882-887 with permission from Elsevier.

The temperature of the leaf might be used... as an index of the condition of the stomata

Francis Darwin 1848 - 1925

Slide courtesy of A. Hetherington

The triple mutant is ABA insensitive in its germination response

Fujii, H., and Zhu, J.-K. (2009). Arabidopsis mutant deficient in 3 abscisic acid-activated protein kinases reveals critical roles in growth, reproduction, and stress. Proc. Natl. Acad. Sci. USA 106: 8380-8385.

There are 34 CDPKs in Arabidopsis. A few have been confirmed to transduce the ABA signal

CDPK

ABA RESPONSES WT cpk4 cpk11 CPK4-over- WT expression Less CPK activity, More CPK activity, Drought tolerance Less drought tolerant More drought tolerant

Zhu, S.-Y., Yu, X.-C., Wang, X.-J., Zhao, R., Li, Y., Fan, R.-C., Shang, Y., Du, S.-Y., Wang, X.-F., Wu, F.-Q., Xu, Y.-H., Zhang, X.-Y., and Zhang, D.-P. (2007). Two calcium-dependent protein kinases, CPK4 and CPK11, regulate abscisic acid signal transduction in Arabidopsis. Plant Cell 19: 3019-3036.

Reprinted from Umezawa, T., Nakashima, K., Miyakawa, T., Kuromori, T., Tanokura, M., Shinozaki, K., and Yamaguchi-Shinozaki, K. (2010). Molecular basis of the core regulatory network in ABA responses: Sensing, signaling and transport. Plant Cell Physiol. 51: 1821-1839 with permission from the Japanese Society of Plant Physiologists.

PP2C

SnRK2 P Some TFs were identified biochemically SnRK2

CDPK P TF

ABA RESPONSES

These transcription factors are highly expressed in seeds, and bind to the RY DNA element (CATGCA(TG)) that is enriched in the promoters of seed-expressed genes

McCarty, D.R., Carson, C.B., Stinard, P.S., and Robertson, D.S. (1989) Molecular analysis of viviparous-1: An abscisic acid-insensitive mutant of maize. Plant Cell 1: 523-532 Giraudat, J., Hauge, B.M., Valon, C., Smalle, J., Parcy, F., and Goodman, H.M. (1992). Isolation of the Arabidopsis ABI3 Gene by positional cloning. Plant Cell 4: 1251-1261. Mönke, G., Altschmied, L., Tewes, A., Reidt, W., Mock, H.-P., Bäumlein, H., and Conrad, U. (2004). Seed-specific transcription factors ABI3 and FUS3: molecular interaction with DNA. Planta 219: 158-166.

Basic Region Leucine Zipper (DNA binding) (Dimerization)

A bZIP protein binding to DNA

Reprinted from Jakoby, M., Weisshaar, B., Dröge-Laser, W., Vicente-Carbajosa, J., Tiedemann, J., Kroj, T., and Parcy, F. (2002). bZIP transcription factors in Arabidopsis. Trends Plant Sci. 7: 106-111 with permission from Elsevier.

Early studies of seed-specific and ABA- responsive genes identified a conserved DNA promoter sequence called the ABA- responsive element (ABRE)

Proteins that bind to the ABRE were identified by gel mobility shift assays and DNase footprinting

These ABRE-binding proteins are called ABFs or AREBS

Mundy, J., Yamaguchi-Shinozaki, K., and Chua, N.H. (1990). Nuclear proteins bind conserved elements in the abscisic acid-responsive promoter of a rice rab gene. Proc. Natl. Acad. Sci. USA 87: 1406-1410. Choi, H.-i., Hong, J.-h., Ha, J.-o., Kang, J.-y., and Kim, S.Y. (2000). ABFs, a family of ABA-responsive element binding factors. J. Biol. Chem. 275: 1723-1730.

ABI5 is a bZIP transcription factor.

Finkelstein, R.R., and Lynch, T.J. (2000). The Arabidopsis abscisic acid response gene ABI5 encodes a basic leucine zipper transcription factor. Plant Cell 12: 599-610, Finkelstein, R.R. (1994). Mutations at two new Arabidopsis ABA response loci are similar to the abi3 mutations. Plant J. 5: 765-771.

Fujita, Y., Fujita, M., Satoh, R., Maruyama, K., Parvez, M.M., Seki, M., Hiratsu, K., Ohme-Takagi, M., Shinozaki, K., and Yamaguchi-Shinozaki, K. (2005). AREB1 is a transcription activator of novel ABRE-dependent ABA signaling that enhances drought stress tolerance in Arabidopsis. Plant Cell 17: 3470-3488.

Overexpression of the ABF3 or ABF4 transcription factors confers drought tolerance

Overexpression of the ABF3 or ABF4 transcription factors causes hyper- sensitive germination

Kang, J.-y., Choi, H.-i., Im, M.-y., and Kim, S.Y. (2002). Arabidopsis basic leucine zipper proteins that mediate stress- responsive abscisic acid signaling. Plant Cell 14: 343-357.

ABA SnRK2 TF

PYR1

SnRK2

PP2C TF

Using protoplasts as an assay system, expression of a TF (ABF2) and SnRK2 is sufficient for ABA-induced gene expression. Also adding PYR1 and ABI1 works too

Reprinted by permission from Macmillan Publishers Ltd: [ Fujii, H., Chinnusamy, V., Rodrigues, A., Rubio, S., Antoni, R., Park, S.-Y., Cutler, S.R., Sheen, J., Rodriguez, P.L., and Zhu, J.-K. (2009). In vitro reconstitution of an abscisic acid signalling pathway. Nature 462: 660-664 copyright 2009.

León, P., Gregorio, J. and Cordoba, E. (2013) ABI4 and its role in chloroplast retrograde communication. Front Plant Sci. 3: 304.

Stress and dehydration - Signaling genes induced genes

A major output of ABA signaling is changes in transcription patterns. Many of the transcriptionally upregulated genes have functions in osmoprotection

Seed- Genes involved in specific genes ABA metabolism

Core Signaling Pathway Mg-chelatase H subunit (CHLH)

Zhang, X.L., Jiang, L., Xin, Q., Liu, Y., Tan, J.X. and Chen, Z.Z. (2015). Structural basis and functions of abscisic acid receptors PYLs. Front. Plant Sci. 6: 88; see also Cutler, S.R., Rodriguez, P.L., Finkelstein, R.R. and Abrams, S.R. (2010). Abscisic Acid: Emergence of a Core Signaling Network. Annu. Rev. Plant Biol. 61: 651-679.

In the absence of ABA, In the presence of WRKY transcription ABA, WRKY factors that are weakly transcription bound by Mg- factors are tightly chelatase subunit H bound at the (CHLH) move to the plastid membrane nucleus and repress and can no longer ABA-responsive genes repress nuclear genes

[The name CHLH derives from the fact that it supports chlorophyll (CHL synthesis by delivering Mg)]

Finkelstein, R. (2013) Abscisic acid synthesis and response. Arabidopsis Book 11: e0166; see also Du, S.Y., Zhang, X.F., Lu, Z., Xin, Q., Wu, Z., Jiang, T., Lu, Y., Wang, X.F. and Zhang, D.P. (2012). Roles of the different components of magnesium chelatase in abscisic acid signal transduction. Plant Mol. Biol. 80: 519–537.

PYR1 PYR/RCAR receptors The core ABA signaling pathway Phosphatase PP2C Protein phosphatases (including ABI1)

Protein kinases (including Kinase SnRK2s and CDPKs)

P P TF

ABA RESPONSES

Turgid guard cells Flaccid guard cells = open stomata = closed stomata = gas exchange = decreased gas exchange + transpiration + decreased transpiration

CO C2

Sirichandra, C., Wasilewska, A., Vlad, F., Valon, C., and Leung, J. (2009a). The guard cell as a single-cell model towards understanding drought tolerance and abscisic acid action. J. Exp. Bot. 60: 1439-1463. by permission of Oxford University Press.

As leaves develop and mature, the ABA concentration declines and older leaves export ABA to younger ones. At the same time, the sensitivity of stomata to ABA increases throughout leaf development

Reprinted from Chater, C.C., Oliver, J., Casson, S. and Gray, J.E. (2014) Putting the brakes on: abscisic acid as a central environmental regulator of stomatal development. New Phytol. 202: 376–391 by permission.

Guard cells responding to ABA Image courtesy Yizhou Wang, University of Glasgow

INNER WALL OPEN

Stomatal movements are mediated by cell turgor, which is controlled by the transport of ions across membranes

PLASMA MEMBRANE / OUTER WALL

INNER WALL OPEN

Ion channels are found on the vacuolar and plasma membranes and include inward and outward transporting K+ channels, anion (A-) channels and Ca2+ channels

PLASMA MEMBRANE / OUTER WALL

H2O OPEN + + + - H O K+ K K K A 2 A- - A- A + A- H2O K ABA causes ion channels in the + H O K 2 vacuolar and plasma membranes to open, releasing ions from the cell, and H2O K+ H2O inactivates a proton-ATPase (red) K+ - A - K+ A H2O H2O K+ H O K+ + A- 2 K

+ + K - K - H2O A H2O A

Water follows

by osmosis H2O H2O H2O

+ + - + + + - K K A A- K K K A

+ + K - K - H2O A H2O A CLOSING

H2O H2O H2O

K+ K+ + A- K

The cell volume shrinks, closing the stomatal pore

The proton pump is reactivated, Potassium ions CLOSED generating a proton gradient and and anions are hyperpolarizing the membrane taken up again H+ H+

K+ + H+ K A- H+

K+ + A- + - K+ K K A A- A- H+ + H+ K

H2O + + + K K - H + A H2O H H2O Water follows by osmosis

© 2015 American Society of Plant Biologists INNER WALL OPEN + K + A- + K+ K K A- - A- The volume of the cell A H O + 2 increases and the K stomatal pore opens K+ H2O H2O + H2O A- + - K+ K K A A- A- H O 2 K+ H2O

H2O

H2O H2O

2+ [Ca ]cyt (nM) ABA

calcium No ABA control

Stomatal aperture

McAinsh, M.R., Brownlee, C., and Hetherington, A.M. (1990). Abscisic acid-induced elevation of guard cell cytosolic Ca2+ precedes stomatal closure. Nature 343: 186- 188; McAinsh, M.R., Brownlee, C., and Hetherington, A.M. (1992). Visualizing changes in cytosolic-free Ca2+ during the response of stomatal guard cells to abscisic acid. Plant Cell 4: 1113-1122.

INNER WALL OPEN

Cytosolic calcium ion 2+ 2+ [Ca ]cyt is a second Ca messenger

Gilroy, S., Read, N.D., and Trewavas, A.J. (1990). Elevation of cytoplasmic calcium by caged calcium or caged inositol trisphosphate initiates stomatal closure. Nature 346: 769-771; Schroeder, J.I., and Hagiwara, S. (1989). Cytosolic calcium regulates ion channels in the plasma membrane of Vicia faba guard cells. Nature 338: 427-430.

© 2015 American Society of Plant Biologists ABA stimulates an increase in 2+ [Ca ]cyt ABA INNER WALL OPEN ABA stimulates an increase in cytoplasmic calcium by activating calcium channels Ca2+ at the plasma and internal membranes

© 2015 American Society of Plant Biologists Membrane voltage and ABA interact 2+ to facilitate [Ca ]cyt increases Before During – During – During – After 2 seconds 5 seconds 15 seconds Membrane hyperpolarization triggers a wave of Ca2+ inwards from the cell periphery.

ABA changes the membrane voltage 2+ threshold at which [Ca ]cyt increases

Grabov, A., and Blatt, M.R. (1998). Membrane voltage initiates Ca2+ waves and potentiates Ca2+ increases with abscisic acid in stomatal guard cells. Proc. Natl. Acad. Sci. USA 95: 4778-4783.

© 2015 American Society of Plant Biologists 2+ Increased [Ca ]cyt helps coordinate the action of many ion channels ABA INNER WALL OPEN Calcium activates ion channels, probably through calcium-dependent protein Ca2+ kinases (CDPKs)

K+

A-

ABA-induced stomatal Anion channel closure is impaired in currents are impaired cpk mutants in cpk mutants

Anion channel activity is reduced in cpk3-1 cpk6-1 mutants as compared to wild-type plants

Mori, IC., Murata, Y., Yang, Y., Munemasa, S, Wang, Y-F., Andreoli, S., Tiriac, H., Alonso, J.M., Harper, J.F., Ecker, J.R., Kwak,, J.M., and Schroeder, J.I. (2006) CDPKs CPK6 and CPK3 function in ABA regulation of guard cell S-type anion- and Ca2+- permeable channels and stomatal closure. PLoS Biol 4(10): e327.

The movement of anions out of the cell increases the pH and depolarizes the membrane, which activates the pH- and voltage-sensitive potassium channels

K+ K+ + A- K

Ca2+ ABA

Pei, Z.-M., Murata, Y., Benning, G., Thomine, S., Klusener, B., Allen, G.J., Grill, E., and Schroeder, J.I. (2000). Calcium channels activated by hydrogen peroxide mediate abscisic acid signalling in guard cells. Nature 406: 731-734.

Dominant abi1 mutant

An inactive form of SnRK2 (labeled with The open stomata caused GFP) was introduced into the right cell of by abi1 can be rescued by a pair of guard cells. The left cell a phosphatase inhibitor responded to ABA and lost turgor; the right cell did not

Reprinted from Li, J., Wang, X.-Q., Watson, M.B., and Assmann, S.M. (2000). Regulation of abscisic acid-induced stomatal closure and anion channels by guard cell AAPK kinase. Science 287: 300-303 with permission from AAAS. Armstrong, F., Leung, J., Grabov, A., Brearley, J., Giraudat, J., and Blatt, M.R. (1995). Sensitivity to abscisic acid of guard-cell K+ channels is suppressed by abi1-1, a mutant Arabidopsis gene encoding a putative protein phosphatase. Proc. Natl. Acad. Sci. USA 92: 9520-9524.

© 2015 American Society of Plant Biologists ABA control of guard cell turgor - summary ABA Guard cell turgor is regulated by a complex network of interacting INNER WALL second messengers, pH, membrane potential, protein phosphorylation, NO ion channel activity – and more!! H2O2 PP2C Ca2+ CDPK SnRK2 K+

K+ K+ + A- K

•Root growth •Vegetative dehydration responses and osmoprotectants •Fruit ripening •Seed development •Biotic stress responses •Drought-tolerant plants

Days post- anthesis Wild- type

ABA↑

Increasing water stress

Sharp, R.E., Silk, W.K., and Hsiao, T.C. (1988). Growth of the maize primary root at low water potentials : I. Spatial distribution of expansive growth. Plant Physiol. 87: 50-57.

Drought stress suppresses lateral root growth while Lateral roots primary root resume growth elongation is upon rewatering maintained

Salt stress induces a prolonged quiescent stage in lateral roots, delaying their outgrowth

ABA synthesis in the endodermis suppresses lateral root outgrowth – perhaps this prevents them from emerging from the primary root into salt-contaminated soil?

Duan, L., Dietrich, D., Ng, C.H., Chan, P.M.Y., Bhalerao, R., Bennett, M.J. and Dinneny, J.R. (2013). Endodermal ABA signaling promotes lateral root quiescence during salt stress in Arabidopsis seedlings. Plant Cell. 25: 324-341.

Well watered

Drought stressed Fluridone (FLU) an ABA synthesis inhibitor interferes with root elongation under drought stress

Spollen, W.G., LeNoble, M.E., Samuels, T.D., Bernstein, N., and Sharp, R.E. (2000). Abscisic acid accumulation maintains maize primary root elongation at low water potentials by restricting ethylene production. Plant Physiol. 122: 967-976. Saab, I.N., Sharp, R.E., Pritchard, J., and Voetberg, G.S. (1990). Increased endogenous abscisic acid maintains primary root growth and inhibits shoot growth of maize seedlings at low water potentials. Plant Physiol. 93: 1329-1336.

Artemia brine shrimp embryos can survive extreme desiccation. They have been sold as pets that “come to life” when put into water

Studying how Artemia survive desiccation is helping us learn how to make human cells desiccation tolerant

© 2015 American Society of Plant Biologists Some plants can tolerate extreme desiccation Watered control A few plants, such as these “resurrection plants” can stay alive even when 90% of their water content is lost

Water withheld 5 days Rewatered Studies of desiccation Rewatered tolerant plants contributes to our understanding of cellular desiccation responses

Selaginella tamariscina Craterostigma plantagineum See also TTPB28:How plants manage water deficit and why it matters

Liu, M.-S., Chien, C.-T., and Lin, T.-P. (2008). Constitutive Components and Induced Gene Expression are Involved in the Desiccation Tolerance of Selaginella tamariscina. Plant and Cell Physiology 49: 653-663, by permission of the Japanese Society of Plant Physiologists; Bohnert, H.J. (2000). What makes desiccation tolerable? Genome Biology, published by BioMed Central.

Osmoprotectants Chaperone functions (sugars, proline, (HSPs, LEAs) glycine betaine)

Movement of Oxidative stress water and ions defense – peroxidase, (aquaporins, ion superoxide dismutase channels)

Reactive oxygen species (ROS) and calcium propagate stress signals. Stomata respond to many cues and integrate local and systemic signals

Suzuki, N., Miller, G., Salazar, C., Mondal, H.A., Shulaev, E., Cortes, D.F., Shuman, J.L., Luo, X., Shah, J., Schlauch, K., Shulaev, V. and Mittler, R. (2013) Temporal-spatial interaction between reactive oxygen species and abscisic acid regulates rapid systemic acclimation in plants. Plant Cell 25: 3553–3569.; Mittler, R. and Blumwald, E. (2015). The roles of ROS and ABA in systemic acquired acclimation. Plant Cell. 27: 64-70.

In many fruits, a peak of ABA synthesis, which often precedes the peak in ethylene synthesis, occurs during ripening or the onset of ripening

Although ripening of all fruits depends on ethylene and ABA, ABA plays a more dominant role in non- climacteric fruits such Relative importance of ABA vs ethylene as grape and citrus

Leng, P., Yuan, B., Guo, Y. and Chen, P. (2014) The role of abscisic acid in fruit ripening and responses to abiotic stress. J. Exp. Bot. 65: 4577–4588; McAtee, P., Karim, S., Schaffer, R. and David, K. (2013) A dynamic interplay between phytohormones is required for fruit development, maturation, and ripening. Front Plant Sci. 4: 79.

© 2015 American Society of Plant Biologists ABA regulates fruit ripening in tomato The zinc-finger transcription factor ZFP2 in tomato represses ABA biosynthesis genes to fine-tune ABA levels during fruit ripening

Days post- anthesis Wild type A lower ABA concentration leads to SlZFP2 over- delayed fruit ripening expression

Days post- anthesis

Wild type An increased ABA concentration leads to SlZFP2 down- more rapid ripening regulation

Weng, L., Zhao, F., Li, R., Xu, C., Chen, K. and Xiao, H. (2015) The zinc finger transcription factor SlZFP2 negatively regulates abscisic acid biosynthesis and fruit ripening in tomato. Plant Physiol. 167: 931–949.

© 2015 American Society of Plant Biologists ABA controls seed maturation, dormancy and desiccation Seed dormancy Germination and desiccation involves catabolism tolerance is GA ABA of ABA and correlated with synthesis of GA high levels of ABA synthesis and accumulation

Embryonic patterning

Reserve Reserve accumulation mobilization

Desiccation tolerance Cell expansion

NO Ethylene synthesis ABA Ethylene synthesis ABA

ABA ABA inactivation Ethylene

ABI3

seed dormancy germination

Arc, E., Sechet, J., Corbineau, F., Rajjou, L. and Marion-Poll, A. (2013) ABA crosstalk with ethylene and nitric oxide in seed dormancy and germination. Front Plant Sci. 4: 63.

Transcription factors work synergistically to control seed development, dormancy and desiccation tolerance

Physical interactions between TFs help confer specificity

Finkelstein, R.R., Gampala, S.S.L., and Rock, C.D. (2002). Abscisic acid signaling in seeds and seedlings. Plant Cell 14: S15-45; Reprinted from Nambara, E., and Marion-Poll, A. (2003). ABA action and interactions in seeds. Trends Plant Sci. 8: 213-217 with permission from Elsevier.

Zea mays vp1 mutant Triple SnRK2 mutant (snrk2.2, 2,3, 2.6)

Nakashima, K., Fujita, Y., Kanamori, N., Katagiri, T., Umezawa, T., Kidokoro, S., Maruyama, K., Yoshida, T., Ishiyama, K., Kobayashi, M., Shinozaki, K., and Yamaguchi-Shinozaki, K. (2009). Three Arabidopsis SnRK2 protein kinases, SRK2D/SnRK2.2, SRK2E/SnRK2.6/OST1 and SRK2I/SnRK2.3, involved in ABA signaling are essential for the control of seed development and dormancy. Plant Cell Physiol. 50: 1345-1363, by permission of the Japanese Society of Plant Biologists.

Water Bacteria Bacteria move Stomata towards close when open bacteria stomata are sensed

Closed stoma Open stoma

Reprinted from Melotto, M., Underwood, W., Koczan, J., Nomura, K., and He, S.Y. (2006). Plant stomata function in innate immunity against bacterial invasion. Cell 126: 969-980 with permission from Elsevier.

Water Bacteria

Pathogens can produce compounds that override the ABA response

Coronatine is a pathogen- produced compound that interferes with ABA-induced stomatal closure

Yi, H., Preuss, M.L., and Jez, J.M. (2009). The devil (and an active jasmonate hormone) is in the details. Nat Chem Biol 5: 273-274. Melotto, M., Underwood, W., Koczan, J., Nomura, K., and He, S.Y. (2006). Plant stomata function in innate immunity against bacterial invasion. Cell 126: 969-980.

ABA enhances some biotic defense responses and interferes with others

Reprinted from Ton, J., Flors, V., and Mauch-Mani, B. (2009). The multifaceted role of ABA in disease resistance. Trends Plant Sci. 14: 310-317 with permission from Elsevier. .

ABA

Defense response This pathogen is more virulent in ABA mutants, This pathogen induces ABA meaning ABA enhances synthesis which makes the resistance plant less resistant Pseudomonas syringae is a Pythium irregulare is a necrotrophic biotrophic pathogen pathogen

Adie, B.A.T., Perez-Perez, J., Perez-Perez, M.M., Godoy, M., Sanchez-Serrano, J.-J., Schmelz, E.A., and Solano, R. (2007). ABA is an essential signal for plant resistance to pathogens affecting JA biosynthesis and the activation of defenses in Arabidopsis. Plant Cell 19: 1665-1681; Reprinted by permission from Macmillan Publishers Ltd. de Torres-Zabala, M., Truman, W., Bennett, M.H., Lafforgue, G., Mansfield, J.W., Rodriguez Egea, P., Bogre, L., and Grant, M. (2007). Pseudomonas syringae pv. tomato hijacks the Arabidopsis abscisic acid signalling pathway to cause disease. EMBO J 26: 1434-1443 copyright 2007.

ABA

Defense response Expression of RD29, an ABA-inducible gene, is diminished in the presence of a pathogen

Reprinted from Kim, T.-H., Hauser, F., Ha, T., Xue, S., Böhmer, M., Nishimura, N., Munemasa, S., Hubbard, K., Peine, N., Lee, B.-h., Lee, S., Robert, N., Parker, Jane E. and Schroeder, Julian I. (2011). Chemical genetics reveals negative regulation of abscisic acid signaling by a plant immune response pathway. Curr. Biol. 21: 990-997 with permission from Elsevier.

Abiotic Biotic stress stress

Brassinosteroids other hormones inhibit ABA Auxin activates responses ABI3 expression eipgenetically via degradation of during early BES1 TPL AXR2/3 seedling HDA19 development via ABI3 and ABI5 ARF Cytokinins promote 10/16 the proteosomal ABI3 degradation of ABI5, and

antagonise the

ABI3 inhibition of Gibberellins germination by ABI5 promote DELLA ABA seed germination Seed germination at high temperature

Climate change, deforestation, population growth are combining to create a “perfect storm” of water scarcity

Projection of percent of time spent under drought conditions in 2050, relative to 2000

Gornall, J., Betts, R., Burke, E., Clark, R., Camp, J., Willett, K., and Wiltshire, A. Implications of climate change for agricultural productivity in the early twenty-first century. Phil. Trans. Royal Soc. B: 365: 2973-2989.

How does ABA contribute to root development, water uptake, rate of transpiration and cellular desiccation responses?

•Modification of ABA synthesis and inactivation to reduce transpiration

•Increased ABA sensitivity of guard cells to reduce transpiration

•Increase root growth for better water uptake

•Drought-inducible expression of desiccation tolerance genes

See also TTPB28:How plants manage water deficit and why it matters

The hormone ABA and its signaling pathway were instrumental in the evolution of land plants

ABA participates in physiological, developmental and defense responses throughout the plant body

Studying ABA is important for the development of drought-tolerant crops

What controls the expression of ABA biosynthesis genes?

How are ABA deactivation and How do ABA transporters conjugation / contribute to ABA deconjugation movement? regulated?

What are the relative contributions of locally synthesized and transported ABA in root and shoot responses?

Reprinted from Nambara, E., and Marion-Pol, A. (2003) ABA action and interactions in seeds. Trends Plant Sci. 8: 213-217 with permission from Elsevier.

© 2015 American Society of Plant Biologists Conclusions and future directions – Signaling PYR1 What is the role of each of the PYR/ RCARs as well as other ABA receptors? What other factors activate SnRK2s? Are their additional PP2C upstream kinases?

How are Ca2+ signals What are other targets for and CDPK activities Ca2+ P SnRK2 phosphorylation? integrated into the P signaling network? SnRK2 How do the CDPK ABA-induced TFs coordinate P their actions? P P How do ROS and NO TF integrate with other signals? Ion H O 2 2 channel NO ABA RESPONSES

ABA RESPONSES Osmoprotectants Chaperone functions

How do guard cells integrate multiple signaling pathways? Water and ion movement Oxidative stress detoxification Why does expression of stress-responsive genes lead to reduced growth rates and yields?

What controls Can we breed plants ion channel that are drought- activities? How do seeds tolerant and integrate ABA and pathogen resistant? other signals?