NHX1 and eIF4A1-stacked transgenic sweetpotato shows enhanced tolerance to drought stress

  • Yandi Zhang
  • Gaifang Deng
  • Weijuan Fan
  • Ling Yuan
  • Hongxia WangEmail author
  • Peng ZhangEmail author
Original Article


Key message

Co-expression of Na+/H+ antiporter NHX1 and DEAD-box RNA helicase eIF4A1 from Arabidopsis positively regulates drought stress tolerance by improving ROS scavenging capacity and maintaining membrane integrity in sweetpotato.


Plants evolve multiple strategies for stress adaptation in nature. To improve sweetpotato resistance to drought stress, transgenic sweetpotato plants overexpressing the Arabidopsis Na+/H+ antiporter, NHX1, and the translation initiation factor elF4A1 were characterized for phenotypic traits and physiological performance. Without drought treatment, the NHX1elF4A1 stacked lines (NE lines) showed normal, vigorous growth comparable to the WT plants. The NE plants showed dense green foliage with delayed leaf senescence and developed more roots than WT plants under drought treatment for 18 days. Compared to WT plants, higher level of reactive oxygen scavenging capacity was detected in NE lines as indicated by reduced H2O2 accumulation as well as increased superoxide dismutase activity and proline content. The relative ion leakage and malondialdehyde content were reduced in NE plants, indicating improved maintenance of intact membranes system. Both NE plants and NHX1-overexpressing plants (N lines) showed larger aerial parts and well-developed root system compared to WT plants under the drought stress conditions, likely due to the improved antioxidant capacity. The NE plants showed better ROS scavenging than N-line plants. All N- and NE-line plants produced normal storage roots with similar yields as WT in the field under normal growth conditions. These results demonstrated the potential to enhance sweetpotato productivity through stacking genes that are involved in ion compartmentalization and translation initiation.


Sweetpotato Drought stress NHX1 eIF4A1 Gene-stacking Overexpression Physiological parameter Phenotype 





Reactive oxygen species


Quantitative reverse transcription-polymerase chain reaction



This work was supported by the grants from the National Key R&D Program of China (2018YFD1000700, 2018YFD1000705), the National Natural Science Foundation of China (31771854) and China Scholarship Council (201804910055). We thank Prof. Sitakanta Pattanaik from UK for corrections the manuscript.

Author contribution statement

YZ analyzed the data and drafted the manuscript. GD performed the majority of the experiments. WF generated the NE transgenic lines. LY revised the manuscript and provided helpful suggestions. HW and PZ coordinated and designed the study and revised the manuscript.

Compliance with ethical standards

Conflict of interest

The authors declare no conflict of interest.

Supplementary material

299_2019_2454_MOESM1_ESM.docx (370 kb)
Supplementary material 1 (DOCX 370 kb)


  1. Abdelkrim YZ, Harigua SE, Barhoumi M, Banroques J, Blondel A, Guizani I, Tanner NK (2018) The steroid derivative 6-aminocholestanol inhibits the DEAD-box helicase eIF4A (LieIF4A) from the Trypanosomatid parasite Leishmania by perturbing the RNA and ATP binding sites. Mol Biochem Parasitol 226:9–19CrossRefPubMedGoogle Scholar
  2. Amin M, Elias SM, Hossain A, Ferdousi A, Rahman MS, Tuteja N, Seraj ZI (2012) Over-expression of a DEAD-box helicase, PDH45, confers both seedling and reproductive stage salinity tolerance to rice (Oryza sativa L.). Mol breed 30(1):345–354CrossRefGoogle Scholar
  3. Apel K, Hirt H (2004) Reactive oxygen species: metabolism, oxidative stress, and signal transduction. Annu Rev Plant Biol 55:373–399CrossRefGoogle Scholar
  4. Apse MP, Aharon GS (1999) Salt tolerance conferred by overexpression of a vacuolar Na+/H+ antiporter in Arabidopsis. Science 285:1256–1258CrossRefPubMedGoogle Scholar
  5. Bartels D, Sunkar R (2005) Drought and salt tolerance in plants. Crit Rev Plant Sci 24:23–58CrossRefGoogle Scholar
  6. Baruah I, Debbarma J, Boruah H, Keshavaiah C (2017) The DEAD-box RNA helicases and multiple abiotic stresses in plants: a systematic review of recent advances and challenges. Plant Omics 10(5):252–262CrossRefGoogle Scholar
  7. Bray EA (1997) Plant responses to water deficit. Trends Plant Sci 2(2):48–54CrossRefGoogle Scholar
  8. Bush MS, Crowe N, Zheng T, Doonan JH (2015) The RNA helicase, eIF 4A-1, is required for ovule development and cell size homeostasis in Arabidopsis. Plant J 84(5):989–1004CrossRefPubMedPubMedCentralGoogle Scholar
  9. Chen H, Jiang JG (2010) Osmotic adjustment and plant adaptation to environmental changes related to drought and salinity. Environ Rev 18:309–319CrossRefGoogle Scholar
  10. Cordin O, Banroques J, Tanner NK, Linder P (2006) The DEAD-box protein family of RNA helicases. Gene 367:17–37CrossRefPubMedGoogle Scholar
  11. Cruz de Carvalho MH (2008) Drought stress and reactive oxygen species: production, scavenging and signaling. Plant Signal Behavior 3(3):156–165CrossRefGoogle Scholar
  12. Dhindsa RS, Matowe W (1989) Drought tolerance in two masses: correlated with enzymatic defense against lipid peroxidation. J Exp Bot 32(126):79–91Google Scholar
  13. Fahad S, Bajwa AA, Nazir U, Anjum SA, Farooq A, Zohaib A, Sadia S, Nasim W, Adkins S, Saud S, Ihsan MZ, Alharby H, Wu C, Wang D, Huang J (2017) Crop production under drought and heat stress: plant responses and management options. Front Plant Sci 8:1147CrossRefPubMedPubMedCentralGoogle Scholar
  14. Fan WJ, Zhang M, Zhang HX, Zhang P (2012) Improved tolerance to various abiotic stresses in transgenic sweet potato (Ipomoea batatas) expressing spinach betaine aldehyde dehydrogenase. PLoS One 7(5):e37344CrossRefPubMedPubMedCentralGoogle Scholar
  15. Fan WJ, Deng GF, Wang HX, Zhang H, Zhang P (2015) Elevated compartmentalization of Na+ into vacuoles improves salt and cold stress tolerance in sweetpotato (Ipomoea batatas). Physiol Plant 154(4):560–571CrossRefPubMedGoogle Scholar
  16. Fedoroff NV, Battisti DS, Beachy RN, Cooper PJ, Fischhoff DA, Hodges CN, Knauf VC, Lobell D, Mazur BJ, Molden D (2010) Radically rethinking agriculture for the 21st century. Science 327:833–834CrossRefPubMedPubMedCentralGoogle Scholar
  17. Gill SS, Tuteja N (2010) Reactive oxygen species and antioxidant machinery in abiotic stress tolerance in crop plants. Plant Physiol Biochem 48(12):909–930CrossRefGoogle Scholar
  18. Guan Q, Wu J, Zhang Y, Jiang C, Liu R, Chai C, Zhu J (2013) A DEAD box RNA helicase is critical for pre-mRNA splicing, cold-responsive gene regulation, and cold tolerance in Arabidopsis. Plant Cell 25(1):342–356CrossRefPubMedPubMedCentralGoogle Scholar
  19. Hanson AD, Nelsen CE, Pedersen AR, Everson EH (1979) Capacity for proline accumulation during water stress in barley and its implications for breeding for drought resistance. Crop Sci 19(4):489–493CrossRefGoogle Scholar
  20. Hu H, Xiong L (2014) Genetic engineering and breeding of drought resistant crops. Annu Rev Plant Biol 65:715–741CrossRefPubMedGoogle Scholar
  21. Jin R, Kim BH, Ji CY, Kim HS, Li HM, Kwak SS (2017) Overexpressing IbCBF3 increases low temperature and drought stress tolerance in transgenic sweetpotato. Plant Physiol Biochem 118:45–54CrossRefPubMedGoogle Scholar
  22. Kang C, Zhai H, Xue L, Zhao N, He S, Liu Q (2018) A lycopene β-cyclase gene, IbLCYB2, enhances carotenoid contents and abiotic stress tolerance in transgenic sweetpotato. Plant Sci 272:243–254CrossRefPubMedGoogle Scholar
  23. Khan A, Garbelli A, Grossi S, Florentin A, Batelli G, Acuna T, Zolla G, Kaye Y, Paul LK, Zhu J-K, Maga G, Grafi G, Barak S (2014) The Arabidopsis STRESS RESPONSE SUPPRESSOR DEAD-box RNA helicases are nucleolar- and chromocenter-localized proteins that undergo stress-mediated relocalization and are involved in epigenetic gene silencing. Plant J 79(1):28–43CrossRefPubMedGoogle Scholar
  24. Kim SH, Hamada T (2005) Rapid and reliable method of extracting DNA and RNA from sweetpotato, Ipomoea batatas (L). Lam. Biotechnol Lett 27:1841–1845CrossRefPubMedGoogle Scholar
  25. Kim SH, Song WK, Kim YH, Kwon SY, Lee HS, Lee IC, Kwak SS (2009) Characterization of full-length enriched expressed sequence tags of dehydration-treated white fibrous roots of sweetpotato. BMB Rep 42(5):271–276CrossRefPubMedGoogle Scholar
  26. Koroleva OA, Calder G, Pendle AF, Kim SH, Lewandowska D, Simpson CG, Jones IM, Brown JWS, Shaw PJ (2009a) Dynamic behavior of Arabidopsis eIF4A-III, putative core protein of exon junction complex: fast relocation to nucleolus and splicing speckles under hypoxia. Plant Cell 21(5):1592–1606CrossRefPubMedPubMedCentralGoogle Scholar
  27. Koroleva OA, Brown JWS, Shaw PE (2009b) Localization of eIF4A-III in the nucleolus and splicing speckles is an indicator of plant stress. Plant Signal Behav 4(12):1148–1151CrossRefPubMedPubMedCentralGoogle Scholar
  28. Kumar S, Kalita A, Srivastava R, Sahoo L (2017) Co-expression of Arabidopsis NHX1 and bar improves the tolerance to salinity, oxidative stress, and herbicide in transgenic Mungbean. Front Plant Sci 8:1896CrossRefPubMedPubMedCentralGoogle Scholar
  29. Leidi EO, Barragan V, Rubio L, El-Hamdaoui A, Ruiz MT, Cubero B, Fernandez JA, Bressan RA, Hasegawa PM, Quintero FJ, Pardo JM (2010) The AtNHX1 exchanger mediates potassium compartmentation in vacuoles of transgenic tomato. Plant J 61:495–506CrossRefPubMedGoogle Scholar
  30. Lesk C, Rowhani P, Ramankutty N (2016) Influence of extreme weather disasters on global crop production. Nature 529(7584):84–87CrossRefGoogle Scholar
  31. Lim S, Kim YH, Kim SH, Kwon SY, Lee HS, Kim JS, Cho KY, Paek KY, Kwak SS (2007) Enhanced tolerance of transgenic sweetpotato plants that express both CuZnSOD and APX in chloroplasts to methyl viologen-mediated oxidative stress and chilling. Mol Breed 19(3):227–239CrossRefGoogle Scholar
  32. Lin KH, Chao PY, Yang CM, Cheng WC, Lo HF, Chang TR (2006) The effects of flooding and drought stresses on the antioxidant constituents in sweet potato leaves. Bot Stud 47(4):417–426Google Scholar
  33. Linder P, Lasko PF, Ashburner M, Leroy P, Nielsen PJ, Nishi K, Schnier J, Slonimski PP (1989) Birth of the DEAD box. Nature 6203(337):121–122CrossRefGoogle Scholar
  34. Liu SP, Zheng LQ, Xue YH, Zhang Q, Wang L, Shou HX (2010) Overexpression of OsVP1 and OsNHX1 increases tolerance to drought and salinity in Rice. Plant Biol 53:444–452CrossRefGoogle Scholar
  35. Loebenstein G, Thottappilly G (2009) The Sweetpotato. Springer, Berlin, pp 13–26CrossRefGoogle Scholar
  36. Lu YY, Deng XP, Kwak SS (2010) Over expression of CuZn superoxide dismutase (CuZnSOD) and ascorbate peroxidase (APX) in transgenic sweet potato enhances tolerance and recovery from drought stress. Afr J Biotechnol 9(49):8378–8391Google Scholar
  37. Luking A, Stahl U, Schmidt U (1998) The protein family of RNA helicases. Crit Rev Biochem Mol Biol 33(4):259–296CrossRefPubMedGoogle Scholar
  38. Luo Y, Liu YB, Dong YX, Gao XQ, Zhang XS (2009) Expression of a putative alfalfa helicase increases tolerance to abiotic stress in Arabidopsis by enhancing the capacities for ROS scavenging and osmotic adjustment. Plant Physiol 166(4):385–394CrossRefGoogle Scholar
  39. Macovei A, Tuteja N (2012) microRNAs targeting DEAD-box helicases are involved in salinity stress response in rice (Oryza sativa L.). BMC Plant Biol 12(1):183CrossRefPubMedPubMedCentralGoogle Scholar
  40. Moore M, Gossmann N, Dietz KJ (2016) Redox regulation of cytosolic translation in plants. Trends Plant Sci 21(5):388–397CrossRefPubMedGoogle Scholar
  41. Munns R, Tester M (2008) Mechanisms of salinity tolerance. Annu Rev Plant Biol 59:651–681CrossRefGoogle Scholar
  42. Muñoz A, Castellano M (2012) Regulation of translation initiation under abiotic stress conditions in plants: is it a conserved or not so conserved process among eukaryotes? Compar Funct Genom 2012:406357CrossRefGoogle Scholar
  43. Rejeb KB, Abdelly C, Savouré A (2014) How reactive oxygen species and proline face stress together. Plant Physiol Biochem 80:278–284CrossRefPubMedGoogle Scholar
  44. Rizhsky L, Hallak HE, Breusegem FV, Rachmilevitch S, Barr JE, Rodermel S, Inze D, Mittler R (2002) Double antisense plants lacking ascorbate peroxidase and catalase are less sensitive to oxidative stress than single antisense plants lacking ascorbate peroxidase or catalase. Plant J 32(3):329–342CrossRefPubMedGoogle Scholar
  45. Ruan L, Chen L, Chen Y, He J, Zhang W, Gao Z, Zhang Y (2012) Expression of Arabidopsis HOMEODOMAIN GLABROUS 11 enhances tolerance to drought stress in transgenic sweet potato plants. J Plant Biol 55(2):151–158CrossRefGoogle Scholar
  46. Sairam RK, Srivastava GC (2002) Changes in antioxidant activity in sub-cellular fractions of tolerant and susceptible wheat genotypes in response to long term salt stress. Plant Sci 162:897–904CrossRefGoogle Scholar
  47. Sanan N, Pham XH, Sudhir KS, Tuteja KS (2005) Pea DNA helicase 45 over expression in tobacco confers high salinity tolerance without affecting yield. Proc Natl Acad Sci USA 102(2):509–514CrossRefGoogle Scholar
  48. Shen GX, Wei J, Qiu XY, Hu RB, Kuppu S, Auld D, Blumwald E, Gaxiola R, Payton P, Zhang H (2015) Co-overexpression of AVP1 and AtNHX1 in cotton further improves drought and salt tolerance in transgenic cotton plants. Plant Mol Bio Rep 33(2):167–177CrossRefGoogle Scholar
  49. Singha DL, Tuteja N, Boro D, Hazarika GN, Singh S (2017) Heterologous expression of PDH47 confers drought tolerance in indica rice. Plant Cell Tiss Org Cult 130(3):577–589CrossRefGoogle Scholar
  50. Tsokanos FF, Albert MA, Demetriades C, Spirohn K, Boutros M, Teleman AA (2016) eIF4A inactivates TORC1 in response to amino acid starvation. EMBO J 35:1058–1076CrossRefPubMedPubMedCentralGoogle Scholar
  51. Tuteja N, Sahoo RK, Garg B, Tuteja R (2013) OsSUV3 dual helicase functions in salinity stress tolerance by maintaining photosynthesis and antioxidant machinery in rice (Oryza sativa L. cv. IR 64). Plant J 76(1):115–127PubMedGoogle Scholar
  52. Vashisht AA, Tuteja N (2005) Cold stress-induced pea DNA helicase 47 is homologous to eIF4A and inhibited by DNA-interacting ligands. Arch Biochem Siophys 440(1):79–90CrossRefGoogle Scholar
  53. Vashisht AA, Tuteja N (2006) Stress responsive DEAD-box helicases: a new pathway to engineer plant stress tolerance. J Photochem Photobiol B Biol 84(2):150–160CrossRefGoogle Scholar
  54. Vashisht AA, Pradhan A, Tuteja R, Tuteja N (2005) Cold and salinity stress induced pea bipolar pea DNA helicase 47 is involved in protein synthesis 110 and stimulated by phosphorylation with protein kinase. Plant J 44(1):76–87CrossRefPubMedGoogle Scholar
  55. Wang B, Zhai H, He S, Zhang H, Ren Z, Zhang D, Liu Q (2016) A vacuolar Na+/H+ antiporter gene, IbNHX2, enhances salt and drought tolerance in transgenic sweetpotato. Sci Horticult 201:153–166CrossRefGoogle Scholar
  56. Xue ZY, Zhi DY (2004) Enhanced salt tolerance of transgenic wheat (Triticum aestivum L.) expressing a vacuolar Na+/H+ antiporter gene with improved grain yields in saline soils in the field and a reduced level of leaf Na+. Plant Sci 167:849–859CrossRefGoogle Scholar
  57. Yan H, Li Q, Park SC, Wang X, Liu YJ, Zhang YG, Tang W, Kou M, Ma DF (2016) Overexpression of CuZnSOD and APX enhance salt stress tolerance in sweet potato. Plant Physiol Biochem 109:20–27CrossRefPubMedGoogle Scholar
  58. Yang AF, Duan XG (2005) Efficient transformation of beet (Beta vulgaris) and production of plants with improved salt-tolerance. Plant Cell Tissue Org Cult 83:259–270CrossRefGoogle Scholar
  59. Yang J, Bi HP, Fan WJ, Zhang M, Wang HX, Zhang P (2011) Efficient embryogenic suspension culturing and rapid transformation of a range of elite genotypes of sweetpotato (Ipomoea batatas L. Lam.). Plant Sci 181:701–711CrossRefPubMedGoogle Scholar
  60. Yin XY, Yang AF (2004) Production and analysis of transgenic maize with improved salt tolerance by the introduction of AtNHX1 gene. Acta Bot Sin 46(7):854–861Google Scholar
  61. Yooyongwech S, Samphumphuang T, Tisarum R, Theerawitaya C, Cha-um S (2017) Water-deficit tolerance in sweet potato [Ipomoea batatas (L.) Lam.] by foliar application of paclobutrazol: role of soluble sugar and free proline. Front Plant Sci 8:1400CrossRefPubMedPubMedCentralGoogle Scholar
  62. Zhai H, Wang F, Si Z, Huo J, Xing L, An Y, He S, Liu Q (2016) A myo-inositol-1-phosphate synthase gene, IbMIPS1, enhances salt and drought tolerance and stem nematode resistance in transgenic sweet potato. Plant Biotechnol J 14(2):592–602CrossRefPubMedGoogle Scholar
  63. Zhang HX, Blumwald E (2001) Transgenic salt-tolerant tomato plants accumulate salt in foliage but not in fruit. Nat Biotechnol 19:765–768CrossRefPubMedGoogle Scholar
  64. Zhang P, Fan W, Wang H, Wu Y, Zhou W, Yang J (2018) Developing new sweet potato varieties with improved performance. In: Wang-Pruski G (ed) Achieving sustainable cultivation of potatoes volume 1, breeding improved varieties. Burleigh Dodds Science Publishing, CambridgeGoogle Scholar
  65. Zhu JK (2003) Regulation of ion homeostasis under salt stress. Curr Opin Plant Biol 6(5):441–445CrossRefPubMedGoogle Scholar
  66. Zhu JK (2016) Abiotic stress signaling and responses in plants. Cell 167(2):313–324CrossRefPubMedPubMedCentralGoogle Scholar
  67. Zhu M, Chen G, Dong T, Wang L, Zhang J, Zhao Z, Hu Z (2015) SlDEAD31, a putative DEAD-box RNA helicase gene, regulates salt and drought tolerance and stress-related genes in tomato. PLoS One 10(8):e0133849CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  1. 1.National Key Laboratory of Plant Molecular Genetics, CAS Center for Excellence in Molecular Plant Sciences, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological SciencesChinese Academy of ScienceShanghaiChina
  2. 2.University of Chinese Academy of SciencesBeijingChina
  3. 3.Shanghai Key Laboratory of Plant Functional Genomics and Resources, Shanghai Chenshan Plant Science Research CenterChinese Academy of ScienceShanghaiChina
  4. 4.Department of Plant and Soil Sciences and Kentucky Tobacco Research and Development CenterUniversity of KentuckyLexingtonUSA

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