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NO and H2O2 contribute to SO2 toxicity via Ca2+ signaling in Vicia faba guard cells

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NO and H2O2 have been implicated as important signals in biotic and abiotic stress responses of plants to the environment. Previously, we have shown that SO2 exposure increased the levels of NO and H2O2 in plant cells. We hypothesize that, as signaling molecules, NO and H2O2 mediate SO2-caused toxicity. In this paper, we show that SO2 hydrates caused guard cell death in a concentration-dependent manner in the concentration range of 0.25 to 6 mmol L−1, which was associated with elevation of intracellular NO, H2O2, and Ca2+ levels in Vicia faba guard cells. NO donor SNP enhanced SO2 toxicity, while NO scavenger c-PTIO and NO synthesis inhibitors l-NAME and tungstate significantly prevented SO2 toxicity. ROS scavenger ascorbic acid (AsA) and catalase (CAT), Ca2+ chelating agent EGTA, and Ca2+ channel inhibitor LaCl3 also markedly blocked SO2 toxicity. In addition, both c-PTIO and AsA could completely block SO2-induced elevation of intracellular Ca2+ level. Moreover, c-PTIO efficiently blocked SO2-induced H2O2 elevation, and AsA significantly blocked SO2-induced NO elevation. These results indicate that extra NO and H2O2 are produced and accumulated in SO2-treated guard cells, which further activate Ca2+ signaling to mediate SO2 toxicity. Our findings suggest that both NO and H2O2 contribute to SO2 toxicity via Ca2+ signaling.

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  1. Arasimowicz M, Floryszak-Wieczorek J (2007) Nitric oxide as a bioactive signaling molecule in plant stress responses. Plant Sci 172:876–887

  2. Arasimowicz M, Floryszak-Wieczorek J, Deckert J, Rucinska R, Gzyl J, Pawlak S, Abramowski D, Jelonek T, Gwózdz EA (2012) Nitric oxide implication in cadmium-induced programmed cell death in roots and signaling response of yellow lupine plants. Plant Physiol Bioch 58:124–134

  3. Besson-Bard A, Gravot A, Richaud P, Auroy P, Duc C, Gaymard F, Taconnat L, Renou JP, Pugin A, Wendehenne D (2009) Nitric oxide contributes to cadmium toxicity in Arabidopsis by promoting cadmium accumulation in roots and by up-regulating genes related to iron uptake. Plant Physiol 149:1302–1315

  4. Black CR, Black VJ (1979) The effects of low concentrations of sulphur dioxide on stomatal conductance and epidermal cell survival in field bean (Vicia faba L.). J Exp Bot 30:291–298

  5. Breusegem FV, Dat JF (2006) Reactive oxygen species in plant cell death. Plant Physiol 141:384–390

  6. Brüne B (2003) Nitric oxide: NO apoptosis or turning it ON? Cell Death Differ 10:864–869

  7. De Pinto MC, Paradiso A, Leonetti P, De Gara L (2006) Hydrogen peroxide, nitric oxide and cytosolic ascorbate peroxidase at the crossroad between defence and cell death. Plant J 48:784–795

  8. Deltoro VI, Gimeno C, Calatayud A, Barreno E (1999) Effect of SO2 fumigation on photosynthetic CO2 gas exchange, chlorophyll a fluorescence emission and antioxidant enzymes in the lichens Evernia prunastri and Ramalina farinacea. Physiol Plant 105:648–654

  9. Domingos P, Prado AM, Wong A, Gehring C, Feijo JA (2015) Nitric oxide: a multitasked signaling gas in plants. Mol Plant 8:506–520

  10. Fan LM, Zhao Z, Assmann SM (2004) Guard cells: a dynamic signaling model. Curr Opin Plant Biol 7:537–546

  11. Felicitas G, Jörg D, Frank G (2013) Nitric oxide, antioxidants and prooxidants in plant defence responses. Front Plant Sci 4:55–60

  12. Floryszak-Wieczorek J, Arasimowicz-Jelonek M, Izbiańska K (2016) The combined nitrate reductase and nitrite-dependent route of NO synthesis in potato immunity to Phytophthora infestans. Plant Physiol Biochem 108:468–477

  13. Gao CJ, Xing D, Li LL, Zhang LR (2008) Implication of reactive oxygen species and mitochondrial dysfunction in the early stages of plant programmed cell death induced by ultraviolet-C overexposure. Planta 227:755–767

  14. Garcia-Mata C, Gay R, Sokolovski S, Hills A, Lamattina L, Blatt MR (2003) Nitric oxide regulated K+ and Cl channels in guard cells through a subset of abscisic acid-evoked signaling pathways. Proc Natl Acad Sci U S A 100:11116–111121

  15. González A, de los ÁCabrera M, Henríquez MJ, Contreras RA, Morales B, Moenne A (2012) Cross talk among calcium, hydrogen peroxide, and nitric oxide and activation of gene expression involving calmodulins and calcium-dependent protein kinases in Ulva compressa exposed to copper excess. Plant Physiol 158:1451–1462

  16. Hao L, Wang Y, Xu J, Feng SD, Ma CY, Liu C, Xu X, Li GZ, Herbert SJ (2011) Role of endogenous salicylic acid in Arabidopsis response to elevated sulfur dioxide concentration. Biol Plantarum 55:297–304

  17. Hausladen A, Stamler JS (1998) Nitric oxide in plant immunity. Proc Natl Acad Sci U S A 95:10345–10347

  18. Haworth M, Gallagher A, Elliott-Kingston C, Raschi A, Marandola D, McElwain JC (2010) Stomatal index responses of Agrostis canina to CO2 and sulphur dioxide: implications for palaeo-[CO2] using the stomatal proxy. New Phytol 188:845–855

  19. Hetherington AM (2001) Guard cell signaling. Cell 107:711–714

  20. Jeandroz S, Lamotte O, Astier J, Rasul S, Trapet P, Besson-Bard A, Bourque S, Nicolas-Francès V, Ma W, Berkowitz GA, Wendehenne D (2013) There’s more to the picture than meets the eye: nitric oxide cross talk with Ca2+ signaling. Plant Physiol 163:459–470

  21. Jiao J, Zhou B, Zhu X, Gao Z, Liang Y (2013) Fusaric acid induction of programmed cell death modulated through nitric oxide signalling in tobacco suspension cells. Planta 238:727–737

  22. Lamotte O, Gould K, Lecourieux D, Sequeira-Legrand A, Lebrun-Garcia A, Durner J, Pugin A, Wendehenne D (2004) Analysis of nitric oxide signaling functions in tobacco cells challenged by the elicitor cryptogein. Plant Physiol 135:516–529

  23. Lamotte O, Courtois C, Dobrowolska G, Besson A, Pugin A, Wendehenne D (2006) Mechanisms of nitric-oxide-induced increase of free cytosolic Ca2+ concentration in Nicotiana plumbaginifolia cells. Free Radic Biol Med 40:1369–1376

  24. Li L, Yi H (2012) Differential expression of Arabidopsis defense-related genes in response to sulfur dioxide. Chemosphere 87:718–724

  25. Li L, Yi H, Wang L, Li X (2008) Effects of sulfur dioxide on the morphological and physiological biochemical parameters in Arabidopsis thaliana plants. J Agro-Environ Sci 27:525–529

  26. Mori IC, Schroeder JI (2004) Reactive oxygen species activation of plant Ca2+ channels, a signaling mechanism in polar growth, hormone transduction, stress signaling, and hypothetically mechanotransduction. Plant Physiol 135:702–708

  27. Neill SJ, Desikan R, Clarke A (2002) Hydrogen peroxide and nitric oxide as signaling molecules in plants. J Exp Bot 53:1237–1242

  28. Niu L, Liao W (2016) Hydrogen peroxide signaling in plant development and abiotic responses: Front Plant Sci 7(13). DOI: 10.3389/fpls.2016.00230

  29. Pei ZM, Murata Y, Benning G, Thomine S, Klüsener B, Allen GJ, Grill E, Schroeder JI (2000) Calcium channels activated by hydrogen peroxide mediate abscisic acid signaling in guard cells. Nature 406:731–734

  30. Qiao W, Fan LM (2014) Cross-talk between nitric oxide and hydrogen peroxide in plant responses to abiotic stresses. Environ Exp Bot 100:84–93

  31. Rakwal R, Agrawal GK, Kubo A, Yonekura M, Tamogami S, Saji H, Iwahashi H (2003) Defense/stress responses elicited in rice seedlings exposed to the gaseous air pollutant sulfur dioxide. Environ Exp Bot 49:223–225

  32. Reape TJ, McCabe PF (2008) Apoptotic-like programmed cell death in plants. New Phytol 180:13–26

  33. Rodríguez-Serrano M, Bárány I, Prem D, Coronado MJ, Risueno MC, Testillano PS (2012) NO, ROS, and cell death associated with caspase-like activity increase in stress-induced microspore embryogenesis of barley. J Exp Bot 63:2007–2024

  34. 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

  35. Shao HB, Chu LY, Lu ZH, Kang CM (2008) Primary antioxidant free radical scavenging and redox signaling pathways in higher plant cells. Int J Biol Sci 4:8–14

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

  37. Van Der Kooij TAW, De-Kok LJ, Haneklaus S, Schnug E (1997) Uptake and metabolism of sulphur dioxide by Arabidopsis thaliana. New Phytol 135:101–107

  38. Wei A, Fu B, Wang Y, Zhai X, Xin X, Zhang C, Cao D, Zhang X (2015) Involvement of NO and H2O2 in sulfur dioxide induced guard cell apoptosis in Tagetes erecta. Ecotoxicol Envion Saf 114:198–203

  39. Ximénez-Embún MG, Ortego F, Castañera P (2016) Drought-stressed tomato plants trigger bottom-up effects on the invasive Tetranychus evansi. PLoS One 11(1):e0145275

  40. Xu Q, Zhang L (2009) Plant caspase-like proteases in plant programmed cell death. Plant Sig Beh 4:902–904

  41. Yamasaki H, Sakihama Y (2000) Simultaneous production of nitric oxide and peroxynitrite by plant nitrate reductase: in vitro evidence for the NR-dependent formation of active nitrogen species. FEBS Lett 468:89–92

  42. Yi H, Liu J, Zheng K (2005) Effect of sulfur dioxide hydrates on cell cycle, sister chromatid exchange and micronuclei in barley. Ecotoxicol Environ Saf 62:421–426

  43. Yi H, Yin J, Liu X, Jing X, Fan S, Zhang H (2012) Sulfur dioxide induced programmed cell death in Vicia guard cells. Ecotoxicol Environ Saf 78:281–286

  44. Yi H, Liu X, Yi M, Chen G (2014) Dual role of hydrogen peroxide in Arabidopsis guard cells in response to sulfur dioxide. Adv Toxicol 2014(6):41–58

  45. Zhao J, Xue M, Bai H, Yi H (2014) Stomatal movement regulation by nitrate reductase-dependent nitric oxide production in Arabidopsis response to sulfur dioxide. Acta Sci Circumst 34:796–800

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We thank Dr. Hong Wang (Monell Chemical Senses Center, USA) for her careful and critical reading of our manuscript. This study was supported by the National Natural Science Foundation of China (Grant No. 30470318, No. 30870454, and No. 31371868), the Research Fund for the Doctoral Program of Higher Education of China (Grant No. 20070108007 and No. 20121401110007), and the Research Project Supported by Shanxi Scholarship Council of China (Grant No. 2009022 and No. 2012013).

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Correspondence to Huilan Yi.

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The authors declare that they have no conflict of interest.

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Responsible editor: Yi-ping Chen

Electronic supplementary material

Suppl. Fig. 1

I Effect of SO2 hydrates on permeability of the guard cell plasma membrane of V. faba. II Esterase activity in V. faba leaves exposed to SO2 hydrates. Different letters between treatments indicate significant differences (P<0.05). Same letter indicates no significant difference (P>0.05). (DOC 43 kb)

Suppl. Fig. 2

The combination of NO scavenger c-PTIO and H2O2 scavenger AsA blocked SO2-caused death of V. faba guard cells. Different letters between treatments indicate significant differences (P<0.05). Same letter indicates no significant difference (P>0.05). (DOC 28 kb)

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Yi, M., Bai, H., Xue, M. et al. NO and H2O2 contribute to SO2 toxicity via Ca2+ signaling in Vicia faba guard cells. Environ Sci Pollut Res 24, 9437–9446 (2017).

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  • Vicia faba guard cell
  • SO2
  • Cytotoxicity
  • Nitric oxide
  • H2O2
  • Ca2+