H2S is involved in ABA-mediated stomatal movement through MPK4 to alleviate drought stress in Arabidopsis thaliana

  • Xinzhe Du
  • Zhuping Jin
  • Liping Zhang
  • Xin Liu
  • Guangdong Yang
  • Yanxi Pei
Regular Article



Hydrogen sulfide (H2S) is a gaseous signaling molecule that participates in multiple physiological processes in both animals and plants. Mitogen-activated protein kinase (MAPK) is important signaling molecule that links the growth and developmental signals and environment stimuli to cellular responses. In the current study we explored the relationship between H2S and MAPK in drought stress resistance in Arabidopsis.


The quantitative real-time (qRT)-PCR, root tip bending experiment and stomatal aperture assay were used in this paper.


Drought stress activated both H2S biosynthesis and gene expression of MAPKs. The increase in MAPK expression was depressed in lcd/des1, a double mutant of H2S synthesis. Then we selected MPK4 as our target and used mpk4 mutants for further studies. H2S was able to alleviate the drought stress in wild-type (WT) Arabidopsis but not in mpk4 mutants. Meanwhile, H2S-induced stomatal movement was impaired in mpk4 mutants. We then examined the role of H2S and MPK4 in stomatal movements in response to abscisic acid (ABA) and hydrogen peroxide (H2O2). ABA- and H2O2- mediated stomatal movements were impaired in lcd/des1 and mpk4 mutants, and H2S-induced stomatal closure was impaired in slac1–3 mutants.


Our results suggested that MPK4 is important downstream of H2S in the drought stress response and in stomatal movement, and that the H2S-MPK4 cascade is involved in ABA-mediated stomatal movement to regulate the drought stress.


H2MPK4 Drought stress Stomata ABA H2O2 



Hydrogen sulfide


Desulfhydrase 1


L-cysteine desulfhydrase


Mitogen-activated protein kinase


Abscisic acid


Pyrabactin Resistance/Pyrabactin resistance-like/Regulatory Component of ABA Receptor


Protein Phosphatase 2C


SNF1-Related Protein Kinases type 2


Slow Anion Channel-Associated 1


Hydrogen peroxide

1/2 MS

1/2 Murashige and Skoog



This work was funded by a grant from the National Natural Science Foundation of China (grant numbers 31672140 to Jin Z., 31671605 to Pei Y.) We thank Shaowu Xue of Huazhong Agricultural University and John Mundy of Copenhagen University for providing seeds generously. The authors have no conflict of interest to declare.

Supplementary material

11104_2018_3894_MOESM1_ESM.doc (42 kb)
ESM 1 (DOC 42 kb)
11104_2018_3894_Fig10_ESM.png (1.8 mb)
Fig. S1

The genotyping of mutants. a-b: the genotyping of lcd/des1. a is genotyping for LCD, and b is genotyping for DES1. c: the genotyping of mpk4 of Ler background. d: the genoptyping of mpk4 of Col background. mpk4 het means mpk4 heterozygote, mpk4 homo means mpk4 homozygote. e: the genotyping of slac1–3. (PNG 723 kb)

11104_2018_3894_Fig8_ESM.png (155 kb)
Fig. S2

The 4-week-old seedlings of mpk4 of Col background. The homozygous or heterozygous of mpk4 with Col background were indicated in the figure. (PNG 278 kb)

11104_2018_3894_Fig9_ESM.png (2 mb)
Fig. S3

The 4-week-old seedling of WT (left) and mpk4 (right) of Ler background. (PNG 86 kb)


  1. Álvarez C, Calo L, Romero LC, García I, Gotor C (2010) An O-acetylserine(thiol)lyase homolog with L-cysteine desulfhydrase activity regulates cysteine homeostasis in Arabidopsis. Plant Physiol 152:656–669. CrossRefPubMedPubMedCentralGoogle Scholar
  2. Danquah A, Zelicourt AD, Colcombet J, Hirt H (2014) The role of ABA and MAPK signaling pathways in plant abiotic stress responses. Biotechnol Adv 32:40–52. CrossRefPubMedGoogle Scholar
  3. Du X, Jin Z, Liu D, Yang G, Pei Y (2017) Hydrogen sulfide alleviates the cold stress through MPK4 in Arabidopsis thaliana. Plant Physiol Biochem 120:112–119. CrossRefPubMedGoogle Scholar
  4. Gudesblat GE, Iusem ND, Morris PC (2007) Guard cell-specific inhibition of Arabidopsis MPK3 expression causes abnormal stomatal responses to abscisic acid and hydrogen peroxide. New Phytol 173:713–721. CrossRefPubMedGoogle Scholar
  5. Hettenhausen C, Baldwin IT, Wu J (2012) Silencing MPK4 in Nicotiana attenuata enhances photosynthesis and seed production but compromises abscisic acid-induced stomatal closure and guard cell-mediated resistance to Pseudomonas syringae pv tomato DC3000. Plant Physiol 158:759–776. CrossRefPubMedGoogle Scholar
  6. Ichimura K, Mizoguchi T, Yoshida R, Yuasa T, Shinozak K (2000) Various abiotic stresses rapidly activate Arabidopsis MAP kinases ATMPK4 and ATMPK6. Plant J 24:655–665. CrossRefPubMedGoogle Scholar
  7. Ichimura K, Shinozaki K, Tena G, Sheen J, Henry Y, Champion A, Kreis M, Zhang S, Hirt H, Wilson C, Heberle-Bors E, Ellis BE, Morris PC, Innes RW, Ecker JR, Scheel D, Klessig DF, Machida Y, Mundy J, Ohashi Y, Walker JC (2002) Mitogen-activated protein kinase cascades in plants: a new nomenclature. Trends In Plant Sci 7:301–308. CrossRefGoogle Scholar
  8. Jammes F, Song C, Shin D, Munemasa S, Takeda K, Gu D, Cho D, Lee S, Giordo R, Sritubtim S, Leonhardt N, Ellis BE, Murata Y, Kwak JM (2009) MAP kinases MPK9 and MPK12 are preferentially expressed in guard cells and positively regulate ROS-mediated ABA signaling. PNAS 106:20520–20525. CrossRefPubMedGoogle Scholar
  9. Jezek M, Blatt MR (2017) The membrane transport system of the guard cell and its integration for stomatal dynamics. Plant Physiol 174:487–519. CrossRefPubMedPubMedCentralGoogle Scholar
  10. Jin Z, Shen J, Qiao Z, Yang G, Wang R, Pei Y (2011) Hydrogen sulfide improves drought resistance in Arabidopsis thaliana. Biochem Biophys Res Commun 414:481–486. CrossRefPubMedGoogle Scholar
  11. Jin Z, Xue S, Luo Y, Tian B, Fang H, Li H, Pei Y (2013) Hydrogen sulfide interacting with abscisic acid in stomatal regulation responses to drought stress in Arabidopsis. Plant Physiol Biochem 62:41–46. CrossRefPubMedGoogle Scholar
  12. Jin Z, Wang Z, Ma Q, Sun L, Zhang L, Liu Z, Liu D, Hao X, Pei Y (2017) Hydrogen sulfide mediates ion fluxes inducing stomatal closure in response to drought stress in Arabidopsis thaliana. Plant Soil 419:141–152. CrossRefGoogle Scholar
  13. Marten H, Hyun T, Gomi K, Seo S, Hedrich R, Roelfsema MRG (2008) Silencing of NtMPK4 impairs CO2-induced stomatal closure, activation of anion channels and cytosolic Ca2+ signals in Nicotiana tabacum guard cells. Plant J 55:698–708. CrossRefPubMedGoogle Scholar
  14. Miura K, Tada Y (2014) Regulation of water, salinity, and cold stress responses by salicylic acid. Front Plant Sci 5:4. CrossRefPubMedPubMedCentralGoogle Scholar
  15. Mizoguchi T, Irie K, Hirayama T, Hayashida N, Yamaguchi-Shinozaki K, Matsumoto K, Shinozaki K (1996) A gene encoding a mitogen-activated protein kinase kinase kinase is induced simultaneously with genes for a mitogen-activated protein kinase and an S6 ribosomal protein kinase by touch, cold, and water stress in Arabidopsis thaliana. PNAS 93:765–769. CrossRefPubMedGoogle Scholar
  16. Papenbrock J, Riemenschneider A, Kamp A, Schulz-Vogt HN, Schmidt A (2007) Characterization of cysteine-degrading and H2S-releasing enzymes of higher plants - from the field to the test tube and back. Plant Biol 9:582–588. CrossRefPubMedGoogle Scholar
  17. Petersen M, Brodersen P, Naested H, Andreasson E, Lindhart U, Johansen B, Nielsen HB, Lacy M, Austin MJ, Parker JE, Sharma SB, Klessig DF, Martienssen R, Mattsson O, Jensen AB, Mundy J (2000) Arabidopsis MAP kinase 4 negatively regulates systemic acquired resistance. Cell 103:1111–1120. CrossRefPubMedGoogle Scholar
  18. Qiao Z, Jing T, Liu Z, Zhang L, Jin Z, Liu D, Pei Y (2015) H2S acting as a downstream signaling molecule of SA regulates Cd tolerance in Arabidopsis. Plant Soil 393:137–146. CrossRefGoogle Scholar
  19. Rodriguez MCS, Petersen M, Mundy J (2010) Mitogen-activated protein kinase signaling in plants. Annu Rev Plant Biol 61:621–649. CrossRefPubMedGoogle Scholar
  20. Šamajová O, Plíhal O, Al-Yousif M, Hirt H, Šamaj J (2013) Improvement of stress tolerance in plants by genetic manipulation of mitogen-activated protein kinases. Biotechnol Adv 31:118–128. CrossRefPubMedGoogle Scholar
  21. Scuffi D, Álvarez C, Laspina N, Gotor C, Lamattina L, García-Mata C (2014) Hydrogen sulfide generated by L-cysteine desulfhydrase acts upstream of nitric oxide to modulate abscisic acid-dependent stomatal closure. Plant Physiol 166:2065–2076. CrossRefPubMedPubMedCentralGoogle Scholar
  22. Scuffi D, Nietzel T, Di Fino LM, Meyer AJ, Lamattina L, Schwarzländer M, Laxalt AM, García-Mata C (2018) Hydrogen sulfide increases production of NADPH oxidase-dependent hydrogen peroxide and phospholipase D-derived phosphatidic acid in guard cell signaling. Plant Physiol 176:2532–2542. CrossRefPubMedGoogle Scholar
  23. Shen J, Xing T, Yuan H, Liu Z, Jin Z, Zhang L, Pei Y (2013) Hydrogen sulfide improves drought tolerance in Arabidopsis thaliana by microRNA expressions. PLoS One 8:e77047. CrossRefPubMedPubMedCentralGoogle Scholar
  24. Shi H, Ye T, Han N, Bian H, Liu X, Chan Z (2015) Hydrogen sulfide regulates abiotic stress tolerance and biotic stress resistance in Arabidopsis. J Intergr Plant Biol 57:628–640. CrossRefGoogle Scholar
  25. Vahisalu T, Kollist H, Wang YF, Nishimura N, Chan WY, Valerio G, Lamminmäki A, Brosché M, Moldau H, Desikan R, Schroeder JI, Kangasjärvi J (2008) SLAC1 is required for plant guard cell S-type anion channel function in stomatal signalling. Nature 452:487–491. CrossRefPubMedPubMedCentralGoogle Scholar
  26. Wang L, Wan R, Shi Y, Xue S (2016) Hydrogen sulfide activates S-type anion channel via OST1 and Ca2+ modules. Mol Plant 9:489–491. CrossRefPubMedGoogle Scholar
  27. Zelicourt AD, Colcombet J, Hirt H (2016) The role of MAPK modules and ABA during abiotic stress signaling. Trends Plant Sci 21:677–685. CrossRefPubMedGoogle Scholar
  28. Zhao W, Zhang J, Lu Y, Wang R (2001) The vasorelaxant effect of H2S as a novel endogenous gaseous KATP channel opener. EMBO J 20:6008–6016. CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer Nature Switzerland AG 2018

Authors and Affiliations

  1. 1.School of Life ScienceShanxi UniversityTaiyuan CityChina
  2. 2.College of Life ScienceQingdao Agricultural UniversityQingdao CityChina
  3. 3.Department of Chemistry and BiochemistryLaurentian UniversitySudburyCanada

Personalised recommendations