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Calcium-dependent protein kinase CPK9 negatively functions in stomatal abscisic acid signaling by regulating ion channel activity in Arabidopsis

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In this manuscript, we demonstrated the negative role of CPK9 in stomatal ABA signaling, and both CPK9 and CPK33 for accurate guard cell function was explored via cpk9/cpk33 double mutants’ phenotype.

Abstract

Abscisic acid (ABA) can inhibit stomatal opening and promote stomatal closure by regulating ion channel activity in guard cell membranes. As an important second messenger, calcium (Ca2+) is essentially needed in ABA regulation of stomatal movement. Calcium-dependent protein kinases (CDPKs) have been proposed to contribute to central Ca2+ signal transduction in plants. Here, we report the functional characterization of CPK9 in Arabidopsis stomatal ABA signaling. CPK9 had high expression in guard cells and the protein was subcellularly located in the cell membrane. A loss-of-function mutant cpk9 showed a much more sensitive phenotype to ABA regulation of stomatal movement and ion channel activity, while CPK9 overexpression lines had opposite phonotypes. These findings demonstrated the negative role of CPK9 in stomatal ABA signaling. As the closest homolog of CPK33, we also proved that stomatal movement of the cpk9/cpk33 double mutants was more sensitive to ABA than either single mutants. These results revealed the role of CPK9 in guard cells, and the need of both CPK9 and CPK33 for accurate guard cell function.

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References

  • Acharya BR, Jeon BW, Zhang W, Assmann SM (2013) Open stomata 1 (OST1) is limiting in abscisic acid responses of Arabidopsis guard cells. New Phytol 200:1049–1063

    Article  CAS  PubMed  Google Scholar 

  • Almadanim MC, Alexandre BM, Rosa MT, Sapeta H, Leitão AE, Ramalho JC, Lam TT, Negrão S, Abreu IA, Oliveira MM (2017) Rice calcium-dependent protein kinase OsCPK17 targets plasma membrane intrinsic protein and sucrose-phosphate synthase and is required for a proper cold stress response. Plant Cell Environ 40:1197–1213

    Article  CAS  PubMed  Google Scholar 

  • Asano T, Kunieda N, Omura Y, Ibe H, Kawasaki T, Takano M, Sato M, Furuhashi H, Mujin T, Takaiwa F, Wu Cy CY, Tada Y, Satozawa T, Sakamoto M, Shimada H (2002) Rice SPK, a calmodulin-like domain protein kinase, is required for storage product accumulation during seed development: phosphorylation of sucrose synthase is a possible factor. Plant Cell 14:619–628

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Asano T, Tanaka N, Yang G, Hayashi N, Komatsu S (2005) Genome-wide identification of the rice calcium-dependent protein kinase and its closely related kinase gene families: comprehensive analysis of the CDPKs gene family in rice. Plant Cell Physiol 46:356–366

    Article  CAS  PubMed  Google Scholar 

  • Asano T, Hayashi N, Kobayashi M, Aoki N, Miyao A, Mitsuhara I, Ichikawa H, Komatsu S, Hirochika H, Kikuchi S (2012) A rice calcium-dependent protein kinase OsCPK12 oppositely modulates salt-stress tolerance and blast disease resistance. Plant J 69:26–36

    Article  CAS  PubMed  Google Scholar 

  • Benetka W, Mehlmer N, Maurer-Stroh S, Sammer M, Koranda M, Neumüller R, Betschinger J, Knoblich JA, Teige M, Eisenhaber F (2008) Experimental testing of predicted myristoylation targets involved in asymmetric cell division and calcium-dependent signalling. Cell Cycle 7:3709–3719

    Article  CAS  PubMed  Google Scholar 

  • Blatt MR, Armstrong F (1993) K+ channels of stomatal guard cells: abscisic acid-evoked control of the outward rectifier mediated by cytoplasmic pH. Planta 191:330–341

    Article  CAS  Google Scholar 

  • Boudsocq M, Sheen J (2013) CDPKs in immune and stress signaling. Trends Plant Sci 18:30–40

    Article  CAS  PubMed  Google Scholar 

  • Cheng SH, Willmann MR, Chen HC, Sheen J (2002) Calcium signaling through protein kinases. The Arabidopsis calcium-dependent protein kinase gene family. Plant Physiol 129:469–485

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Choi HI, Park HJ, Park JH, Kim S, Im MY, Seo HH, Kim YW, Hwang I, Kim SY (2005) Arabidopsis calcium-dependent protein kinase AtCPK32 interacts with ABF4, a transcriptional regulator of abscisic acid-responsive gene expression, and modulates its activity. Plant Physiol 139:1750–1761

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Clough SJ, Bent AF (1998) Floral dip: a simplified method for Agrobacterium-mediated transformation of Arabidopsis thaliana. Plant J 16:735–743

    Article  CAS  PubMed  Google Scholar 

  • Corratgé-Faillie C, Ronzier E, Sanchez F, Prado K, Kim JH, Lanciano S, Leonhardt N, Lacombe B, Xiong TC (2017) The Arabidopsis guard cell outward potassium channel GORK is regulated by CPK33. FEBS Lett 591:1982–1992

    Article  CAS  PubMed  Google Scholar 

  • Cousson A, Vavasseur A (1998) Putative involvement of cytosolic Ca2+ and GTP-binding proteins in cyclic-GMP-mediated induction of stomatal opening by auxin in Commelina communis L. Planta 206:308–314

    Article  CAS  Google Scholar 

  • Dammann C, Ichida A, Hong B, Romanowsky SM, Hrabak EM, Harmon AC, Pickard BG, Harper JF (2003) Subcellular targeting of nine calcium-dependent protein kinase isoforms from Arabidopsis. Plant Physiol 132:1840–1848

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Dubiella U, Seybold H, Durian G, Komander E, Lassig R, Witte C, Schulze WX, Romeis T (2013) Calcium-dependent protein kinase/NADPH oxidase activation circuit is required for rapid defense signal propagation. Proc Natl Acad Sci USA 110:8744–8749

    Article  PubMed  Google Scholar 

  • Franz S, Ehlert B, Liese A, Kurth J, Cazale A, Romeis T (2011) Calcium-Dependent protein kinase CPK21 functions in abiotic stress response in Arabidopsis thaliana. Mol Plant 4:83–96

    Article  CAS  PubMed  Google Scholar 

  • Gambale F, Uozumi N (2006) Properties of shaker-type potassium channels in higher plants. J Membr Biol 210:1–19

    Article  CAS  PubMed  Google Scholar 

  • Hrabak EM, Chan CWM, Gribskov M, Harper JF, Choi JH, Halford NG, Kudla J, Luan S, Nimmo HG, Sussman MR (2003) The Arabidopsis CDPK-SnRK superfamily of protein kinases. Plant Physiol 132:666–680

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Jiang S, Zhang D, Wang L, Pan J, Liu Y, Kong X, Zhou Y, Li D (2013) A maize calcium-dependent protein kinase gene, ZmCPK4, positively regulated abscisic acid signaling and enhanced drought stress tolerance in transgenic Arabidopsis. Plant Physiol Biochem 71:112–120

    Article  CAS  PubMed  Google Scholar 

  • Johnson DR, Bhatnagar RS, Knoll LJ, Gordon JI (1994) Genetic and biochemical studies of protein N-myristoylation. Annu Rev Biochem 63:869–914

    Article  CAS  PubMed  Google Scholar 

  • Lee SC, Lim CW, Lan W, He K, Luan S (2013) ABA signaling in guard cells entails a dynamic protein–protein interaction relay from the PYL-RCAR family receptors to ion channels. Mol Plant 6:528–538

    Article  CAS  PubMed  Google Scholar 

  • Li J, Wang X, Watson MB, Assmann SM (2000) Regulation of abscisic acid-induced stomatal closure and anion channels by guard cell AAPK Kinase. Science 287:300–303

    Article  CAS  PubMed  Google Scholar 

  • Li CL, Wang M, Ma XY, Zhang W (2014) NRGA1, a putative mitochondrial pyruvate carrier, mediates ABA regulation of guard cell ion channels and drought stress responses in Arabidopsis. Mol Plant 7:1508–1521

    Article  CAS  PubMed  Google Scholar 

  • Li CL, Wang M, Wu XM, Chen DH, Lv HJ, Shen JL, Qiao Z, Zhang W (2016) THI1, a thiamine thiazole synthase, interacts with Ca2+-dependent protein kinase CPK33 and modulates the S-type anion channels and stomatal closure in Arabidopsis. Plant Physiol 170:1090–1104

    Article  CAS  PubMed  Google Scholar 

  • Liu K-h, Niu Y, Konishi M, Wu Y, Du H, Chung HS, Li L, Boudsocq M, McCormack M, Maekawa S (2017) Discovery of nitrate–CPK–NLP signalling in central nutrient–growth networks. Nature 545:311

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2– ∆∆Ct method. Methods 25:402–408

    Article  CAS  Google Scholar 

  • Ma S, Wu W (2007) AtCPK23 functions in Arabidopsis responses to drought and salt stresses. Plant Mol Biol 65:511–518

    Article  CAS  PubMed  Google Scholar 

  • McAinsh MR, Webb AAR, Taylor JE, Hetherington AM (1995) Stimulus-induced oscillations in guard cell cytosolic free calcium. Plant Cell 7:1207–1219

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Mehlmer N, Wurzinger B, Stael S, Hofmann-Rodrigues D, Csaszar E, Pfister B, Bayer R, Teige M (2010) The Ca2+-dependent protein kinase CPK3 is required for MAPK-independent salt-stress acclimation in Arabidopsis. Plant J 63:484–498

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Mori IC, Murata Y, Yang Y, Munemasa S, Wang Y, Andreoli S, Tiriac H, Alonso JM, Harper JF, Ecker JR (2006) CDPKs CPK6 and CPK3 function in ABA regulation of guard cell S-type anion- and Ca2+-permeable channels and stomatal closure. PLoS Biol 4:1749–1762

    Article  CAS  Google Scholar 

  • Oliver B, Jorg K (2012) Analysis of calcium signaling pathways in plants. Biochim Biophys Acta 1820:1283–1293

    Article  CAS  Google Scholar 

  • Pandey S, Zhang W, Assmann SM (2007) Roles of ion channels and transporters in guard cell signal transduction. FEBS Lett 581:2325–2336

    Article  CAS  PubMed  Google Scholar 

  • Patharkar OR, Cushman JC (2000) A stress-induced calcium-dependent protein kinase from Mesembryanthemum crystallinum phosphorylates a two-component pseudo-response regulator. Plant J 24:679–691

    Article  CAS  PubMed  Google Scholar 

  • Pei ZM, Kuchitsu K, Ward JM, Schwarz M, Schroeder JI (1997) Differential abscisic acid regulation of guard cell slow anion channels in Arabidopsis wild-type and abi1 and abi2 mutants. Plant Cell 9:409–423

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Schroeder JI, Raschke K, Neher E (1987) Voltage dependence of K+ channels in guard-cell protoplasts. Proc Natl Acad Sci USA 84:4108–4112

    Article  CAS  PubMed  Google Scholar 

  • Schroeder JI, Allen GJ, Hugouvieux V, Kwak JM, Waner D (2001) Guard cell signal transduction. Annu Rev Plant Physiol Plant Mol Biol 52:627–658

    Article  CAS  PubMed  Google Scholar 

  • Schwartz A, Wu WH, Tucker EB, Assmann SM (1994) Inhibition of inward K+ channels and stomatal response by abscisic acid: an intracellular locus of phytohormone action. Proc Natl Acad Sci USA 91:4019–4023

    Article  CAS  PubMed  Google Scholar 

  • Sheen J (1996) Ca2+-dependent protein kinases and stress signal transduction in plants. Science 274:1900–1902

    Article  CAS  PubMed  Google Scholar 

  • Shi S, Li S, Asim M, Mao J, Xu D, Ullah Z, Liu G, Wang Q, Liu H (2018) The Arabidopsis calcium-dependent protein kinases (CDPKs) and their roles in plant growth regulation and abiotic stress responses. Int J Mol Sci 19:1900–1918

    Article  CAS  PubMed Central  Google Scholar 

  • Very AA, Sentenac H (2003) Molecular mechanisms and regulation of K+ transport in higher plants. Annu Rev Plant Biol 54:575–603

    Article  CAS  PubMed  Google Scholar 

  • Wang XQ, Ullah H, Jones AM, Assmann SM (2001) G protein regulation of ion channels and abscisic acid signaling in Arabidopsis guard cells. Science 292:2070–2072

    Article  CAS  PubMed  Google Scholar 

  • Wang L, Yu C, Xu S, Zhu Y, Huang W (2016) OsDi19-4 acts downstream of OsCDPK14 to positively regulate ABA response in rice. Plant Cell Environ 39:2740–2753

    Article  CAS  PubMed  Google Scholar 

  • Wei S, Hu W, Deng X, Zhang Y, Liu X, Zhao X, Luo Q, Jin Z, Li Y, Zhou S (2014) A rice calcium-dependent protein kinase OsCPK9 positively regulates drought stress tolerance and spikelet fertility. BMC Plant Biol 14:133

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Zhang W, Nilson SE, Assmann SM (2008) Isolation and whole-cell patch clamping of Arabidopsis guard cell protoplasts. Cold Spring Harbor Protoc 2008:pdb prot5014

    Article  Google Scholar 

  • Zhu SY, Yu XC, Wang XJ, Zhao R, Li Y, Fan RC, Shang Y, Du SY, Wang XF, Wu FQ, Xu YH, Zhang XY, Zhang DP (2007) Two calcium-dependent protein kinases, CPK4 and CPK11, regulate abscisic acid signal transduction in Arabidopsis. Plant Cell 19:3019–3036

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Zou J, Wei F, Wang C, Wu J, Ratnasekera D, Liu W, Wu W (2010) Arabidopsis calcium-dependent protein kinase CPK10 functions in abscisic acid- and Ca2+-mediated stomatal regulation in response to drought stress. Plant Physiol 154:1232–1243

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Zou J, Li XD, Ratnasekera D, Wang C, Liu WX, Song LF, Zhang WZ, Wu WH (2015) Arabidopsis CALCIUM-DEPENDENT PROTEIN KINASE8 and CATALASE3 function in abscisic acid-mediated signaling and H2O2 homeostasis in stomatal guard cells under drought stress. Plant Cell 27:1445–1460

    Article  CAS  PubMed  PubMed Central  Google Scholar 

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Acknowledgements

This work was supported by National Natural Science Foundation of China (31500211), the START-UP Fund from Jiangsu Normal University (17XLR039) and the Science Fund for Distinguished Young Scholars from Shandong University.

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Author notes

  1. Dong-Hua Chen and Hui-Ping Liu have contributed equally to this work.

Authors and Affiliations

  1. Key Laboratory of Plant Cell Engineering and Germplasm Innovation, Ministry of Education; School of Life Science, Shandong University, Qingdao, 266237, China

    Dong-Hua Chen & Hui-Ping Liu

  2. College of Life Science, Jiangsu Normal University, Xuzhou, 221116, China

    Chun-Long Li

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  1. Dong-Hua Chen
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CL designed the experiments; CL, DC, HL performed the experiments, analyzed and discussed the results; CL and DC wrote the manuscript.

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Correspondence to Chun-Long Li.

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Chen, DH., Liu, HP. & Li, CL. Calcium-dependent protein kinase CPK9 negatively functions in stomatal abscisic acid signaling by regulating ion channel activity in Arabidopsis. Plant Mol Biol 99, 113–122 (2019). https://doi.org/10.1007/s11103-018-0805-y

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