Constitutive expression of an A-5 subgroup member in the DREB transcription factor subfamily from Ammopiptanthus mongolicus enhanced abiotic stress tolerance and anthocyanin accumulation in transgenic Arabidopsis

Autoři: Meiyan Ren aff001;  Zhilin Wang aff001;  Min Xue aff001;  Xuefeng Wang aff001;  Feng Zhang aff001;  Yu Zhang aff001;  Wenjun Zhang aff001;  Maoyan Wang aff001
Působiště autorů: College of Life Sciences, Inner Mongolia Agricultural University, Hohhot, China aff001
Vyšlo v časopise: PLoS ONE 14(10)
Kategorie: Research Article
doi: 10.1371/journal.pone.0224296


Dehydration-responsive element-binding (DREB) transcription factors (TFs) are key regulators of stress-inducible gene expression in plants. Anthocyanins, an important class of flavonoids, protect plants from reactive oxygen species produced under abiotic stresses. However, regulation of DREBs on anthocyanin accumulation is largely unknown. Here, an A-5 subgroup DREB gene (AmDREB3) isolated from Ammopiptanthus mongolicus, a desert broadleaf shrub with very high tolerance to harsh environments, was characterized in terms of both abiotic stress tolerance and anthocyanin accumulation. AmDREB3 does not contain the transcriptional repression motif EAR, and the protein was located in the nucleus and has transcriptional activation capacity. The transcription of AmDREB3 was differentially induced in the shoots and roots of A. mongolicus seedlings under drought, salt, heat, cold, ultraviolet B, and abscisic acid treatments. Moreover, the transcript levels in twigs, young leaves, and roots were higher than in other organs of A. mongolicus shrubs. Constitutively expressing AmDREB3 improved the tolerance of transgenic Arabidopsis to drought, high salinity and heat, likely by inducing the expression of certain stress-inducible genes. The transgenic Arabidopsis seedlings also exhibited an obvious purple coloration and significant increases in anthocyanin accumulation and/or oxidative stress tolerance under drought, salt, and heat stresses. These results suggest that the AmDREB3 TF may be an important positive regulator of both stress tolerance and anthocyanin accumulation.

Klíčová slova:

Arabidopsis thaliana – Drought adaptation – Genetically modified plants – Leaves – Plant resistance to abiotic stress – Polymerase chain reaction – Seedlings – Thermal stresses


1. Wang H, Wang H, Shao H, Tang X. Recent advances in utilizing transcription factors to improve plant abiotic stress tolerance by transgenic technology. Front Plant Sci. 2016; 7:67. doi: 10.3389/fpls.2016.00067 26904044

2. Erpen L, Devi HS, Grosser JW, Dutt M. Potential use of the DREB/ERF, MYB, NAC and WRKY transcription factors to improve abiotic and biotic stress in transgenic plants. Plant Cell Tiss Organ Cult. 2018; 132(1):1–25. doi: 10.1007/s11240-017-1320-6

3. Lata C, Prasad M. Role of DREBs in regulation of abiotic stress responses in plants. J Exp Bot. 2011; 62:4731–4748. doi: 10.1093/jxb/err210 21737415

4. Sakuma Y, Liu Q, Dubouzet JG, Abe H, Shinozaki K, Yamaguchi-Shinozaki K. DNA-binding specificity of the ERF/AP2 domain of Arabidopsis DREBs, transcription factors involved in dehydration- and cold-inducible gene expression. Biochem Biophys Res Commun. 2002; 290:998–1009. doi: 10.1006/bbrc.2001.6299 11798174

5. Agarwal PK, Gupta K, Lopato S, Agarwal P. Dehydration responsive element binding transcription factors and their applications for the engineering of stress tolerance. J Exp Bot. 2017; 68(9):2135–2148. doi: 10.1093/jxb/erx118 28419345

6. Dong CJ, Liu JY. The Arabidopsis EAR-motif-containing protein RAP2.1 functions as an active transcriptional repressor to keep stress responses under tight control. BMC Plant Biol. 2010; 10:47. doi: 10.1186/1471-2229-10-47 20230648

7. Huang B, Liu JY. A cotton dehydration responsive element binding protein functions as a transcriptional repressor of DRE-mediated gene expression. Biochem Biophys Res Commun. 2006; 343(4):1023–31. doi: 10.1016/j.bbrc.2006.03.016 16574068

8. Dong CJ, Huang B, Liu JY. The cotton dehydration-responsive element binding protein GhDBP1 contains an EAR-motif and is involved in the defense response of Arabidopsis to salinity stress. Functional Plant Biology. 2010; 37(1):64–73. doi: 10.1071/FP09100

9. Zhao XJ, Lei HJ, Zhao K, Yuan HZ, Li TH. Isolation and characterization of a dehydration responsive element binding factor MsDREBA5 in Malus sieversii Roem. Scientia horticulturae. 2012; 142:212–220. dio: doi: 10.1016/j.scienta.2012.05.020

10. Jin T, Chang Q, Li W, Yin D, Li Z, Wang D, et al. Stress-inducible expression of GmDREB1 conferred salt tolerance in transgenic alfalfa. Plant Cell Tiss Organ Cult. 2010; 100: 219. doi: 10.1007/s11240-009-9628-5

11. Chen M, Wang QY, Cheng XG, Xu ZS, Li LC, Ye XG, et al. GmDREB2, a soybean DRE-binding transcription factor, conferred drought and high-salt tolerance in transgenic plants. Biochemical and biophysical research communications. 2007; 353(2):299–305. doi: 10.1016/j.bbrc.2006.12.027 17178106

12. Chen M, Xu Z, Xia L, Li L, Cheng X, Dong J, et al. Cold-induced modulation and functional analyses of the DRE-binding transcription factor gene, GmDREB3, in soybean (Glycine max L.). Journal of experimental botany. 2009; 60(1):121–135. doi: 10.1093/jxb/ern269 18988621

13. Bouaziz D, Pirrello J, Ben Amor H, Hammami A, Charfeddine M, Dhieb A, et al. Ectopic expression of dehydration responsive element binding proteins (StDREB2) confers higher tolerance to salt stress in potato. Plant Physiology and Biochemistry. 2012; 60:98–108. doi: 10.1016/j.plaphy.2012.07.029 22922109

14. Zhou ML, Ma JT, Zhao YM, Wei YH, Tang YX, Wu YM. Improvement of drought and salt tolerance in Arabidopsis and Lotus corniculatus by overexpression of a novel DREB transcription factor from Populus euphratica. Gene. 2012; 506(1):10–17. doi: 10.1016/j.gene.2012.06.089 22771912

15. Ma JT, Yin CC, Zhou ML, Wang ZL, Wu YM. A novel DREB transcription factor from Halimodendron halodendron leads to enhance drought and salt tolerance in Arabidopsis. Biologia plantarum. 2015; 59(1):74–82. doi: 10.1007/s10535-014-0467-9

16. Liu N, Zhong NQ, Wang GL, Li LJ, Liu XL, He YK, et al. Cloning and functional characterization of PpDBF1 gene encoding a DRE-binding transcription factor from Physcomitrella patens. Planta. 2007; 226(4):827–838. doi: 10.1007/s00425-007-0529-8 17541631

17. Li H, Zhang D, Li X, Guan K, Yang H. Novel DREB A-5 subgroup transcription factors from desert moss (Syntrichia caninervis) confers multiple abiotic stress tolerance to yeast. Journal of plant physiology. 2016; 194:45–53. doi: 10.1016/j.jplph.2016.02.015 27016184

18. Van den Ende W, El-Esawe S K. Sucrose signaling pathways leading to fructan and anthocyanin accumulation: a dual function in abiotic and biotic stress responses?. Environmental and Experimental Botany. 2014; 108:4–13. doi: 10.1016/j.envexpbot.2013.09.017

19. Mouradov A and Spangenberg G. Flavonoids: a metabolic network mediating plants adaptation to their real estate. Front Plant Sci. 2014; 5:620. doi: 10.3389/fpls.2014.00620 25426130

20. Shi MZ and Xie DY. Biosynthesis and Metabolic Engineering of Anthocyanins in Arabidopsis thaliana. Recent Pat Biotechnol. 2014; 8(1): 47–60. doi: 10.2174/1872208307666131218123538 24354533

21. Xie Y, Chen P, Yan Y, Bao C, Li X, Wang L, et al. An atypical R2R3 MYB transcription factor increases cold hardiness by CBF-dependent and CBF-independent pathways in apple. New Phytologist. 2018; 218(1):201–218. doi: 10.1111/nph.14952 29266327

22. Mahmood K, Xu Z, El-Kereamy A, Casaretto JA, Rothstein SJ. The Arabidopsis transcription factor ANAC032 represses anthocyanin biosynthesis in response to high sucrose and oxidative and abiotic stresses. Frontiers in plant science. 2016: 7:1548. doi: 10.3389/fpls.2016.01548 27790239

23. Borevitz JO, Xia Y, Blount J, Dixon RA, Lamb C. Activation tagging identifies a conserved MYB regulator of phenylpropanoid biosynthesis. Plant Cell. 2000; 12(12):2383–2393. doi: 10.1105/tpc.12.12.2383 11148285

24. Gonzalez A, Zhao M, Leavitt JM and Lloyd AM. Regulation of the anthocyanin biosynthetic pathway by the TTG1/bHLH/Myb transcriptional complex in Arabidopsis seedlings. Plant J. 2008; 53:814–827. doi: 10.1111/j.1365-313X.2007.03373.x 18036197

25. Pesch M, Schultheiss I, Klopffleisch K, Uhrig JF, Koegl M, Clemen CS, et al. TRANSPARENT TESTA GLABRA1 and GLABRA1 compete for binding to GLABRA3 in Arabidopsis. Plant Physiol. 2015; 168: 584–597. doi: 10.1104/pp.15.00328 25926482

26. Gou JY, Felippes FF, Liu CJ, Weigel D, Wang JW. Negative regulation of anthocyanin biosynthesis in Arabidopsis by a miR156-targeted SPL transcription factor. Plant Cell. 2011; 23(4):1512–22. doi: 10.1105/tpc.111.084525 21487097

27. Rajagopalan R, Vaucheret H, Trejo J, Bartel DP. A diverse and evolutionarily fluid set of microRNAs in Arabidopsis thaliana. Genes Dev. 2006; 20(24):3407–25. doi: 10.1101/gad.1476406 17182867

28. Luo QJ, Mittal A, Jia F, Rock CD. An autoregulatory feedback loop involving PAP1 and TAS4 in response to sugars in Arabidopsis. Plant molecular biology. 2012; 80(1):117–129. doi: 10.1007/s11103-011-9778-9 21533841

29. Pino MT, Skinner JS, Jeknić Z, Hayes PM, Soeldner AH, Thomashow MF, et al. Ectopic AtCBF1 overexpression enhances freezing tolerance and induces cold acclimation-associated physiological modifications in potato. Plant cell & environment. 2008; 31(4):393–406. doi: 10.1111/j.1365-3040.2008.01776.x 18182016

30. Navarro M, Ayax C, Martinez Y, Laur J, El Kayal W, Marque C, et al. Two EguCBF1 genes overexpressed in Eucalyptus display a different impact on stress tolerance and plant development. Plant biotechnology journal. 2011; 9(1):50–63. doi: 10.1111/j.1467-7652.2010.00530.x 20492548

31. Wisniewski M, Norelli J, Bassett C, Artlip T, Macarisin D. Ectopic expression of a novel peach (Prunus persica) CBF transcription factor in apple (Malus× domestica) results in short-day induced dormancy and increased cold hardiness. Planta. 2011; 233(5), 971–983. doi: 10.1007/s00425-011-1358-3 21274560

32. Gu X, Chen Y, Gao Z, Qiao Y, Wang X. Transcription factors and anthocyanin genes related to low-temperature tolerance in rd29A: RdreB1BI transgenic strawberry. Plant Physiology and Biochemistry. 2015; 89:31–43. doi: 10.1016/j.plaphy.2015.02.004 25686702

33. An D, Ma Q, Yan W, Zhou W, Liu G, Zhang P. Divergent regulation of CBF regulon on cold tolerance and plant phenotype in cassava overexpressing Arabidopsis CBF3 gene. Frontiers in plant science. 2016; 7:1866. doi: 10.3389/fpls.2016.01866 27999588

34. Artlip TS, Wisniewski ME, Arora R, Norelli JL. An apple rootstock overexpressing a peach CBF gene alters growth and flowering in the scion but does not impact cold hardiness or dormancy. Hortic Res. 2016; 3:16006. doi: 10.1038/hortres.2016.6 26981253

35. Wu Y, Wei W, Pang X, Wang X, Zhang H, Dong B, et al. Comparative transcriptome profiling of a desert evergreen shrub, Ammopiptanthus mongolicus, in response to drought and cold stresses. BMC Genomics. 2014; 15:671. doi: 10.1186/1471-2164-15-671 25108399

36. Gao F, Wang N, Li H, Liu J, Fu C, Xiao Z, et al. Identification of drought-responsive microRNAs and their targets in Ammopiptanthus mongolicus by using high-throughput sequencing. Sci Rep. 2016; 6:34601. doi: 10.1038/srep34601 27698373

37. Yin Y, Jiang X, Ren M, Xue M, Nan D, Wang Z, et al. AmDREB2C, from Ammopiptanthus mongolicus, enhances abiotic stress tolerance and regulates fatty acid composition in transgenic Arabidopsis. Plant Physiol Biochem. 2018; 130: 517–528. doi: 10.1016/j.plaphy.2018.08.002 30096686

38. Thomas DS, Kenneth JL. Analyzing real-time PCR data by the comparative Ct method. Nature Protocols. 2008; 3(6): 1101–1108. doi: 10.1038/nprot.2008.73 18546601

39. Kumar S, Nei M, Dudley J, Tamura K. MEGA: a biologist-centric software for evolutionary analysis of DNA and protein sequences. Brief. Bioinform. 2008; 9: 299–306. doi: 10.1093/bib/bbn017 18417537

40. Yoo SD, Cho YH, Sheen J. Arabidopsis mesophyll protoplasts: a versatile cell system for transient gene expression analysis. Nat Protoc. 2007; 2(7):1565–72. doi: 10.1038/nprot.2007.199 17585298

41. Clough SJ, Bent AF. Floral dip: a simplified method for Agrobacterium-mediated transformation of Arabidopsis thaliana. Plant J. 1998; 16(6):735–743. doi: 10.1046/j.1365-313x.1998.00343.x 10069079

42. Zhang W, Ning G, Lv H, Liao L, Bao M. Single MYB-type transcription factor AtCAPRICE: a new efficient tool to engineer the production of anthocyanin in tobacco. Biochem Biophys Res Commun. 2009; 388(4):742–747. doi: 10.1016/j.bbrc.2009.08.092 19703423

43. Joshi R, Wani SH, Singh B, Bohra A, Dar ZA, Lone AA, et al. Transcription factors and plants response to drought stress: current understanding and future directions. Frontiers in Plant Science. 2016; 7:1029. doi: 10.3389/fpls.2016.01029 27471513

44. Kazan K, Manners JM. MYC2: the master in action. Molecular plant. 2013; 6(3):686–703. doi: 10.1093/mp/sss128 23142764

45. Yamaguchi-Shinozaki K, Kazuo S. 'DREB regulons in abiotic-stress-responsive gene expression in plants.' Molecular Breeding of Forage and Turf. 2009; 15–28. doi: 10.1007/978-0-387-79144-9_2

46. Dubouzet JG, Sakuma Y, Ito Y, Kasuga M, Dubouzet EG, Miura S, et al. OsDREB genes in rice, Oryza sativa L., encode transcription activators that function in drought-, high-salt- and cold-responsive gene expression. Plant J. 2003; 33:751–763. doi: 10.1046/j.1365-313x.2003.01661.x 12609047

47. Zhou ML, Ma JT, Pang JF, Zhang ZL, Tang YX, Wu YM. Regulation of plant stress response by dehydration responsive element binding (DREB) transcription factors. African Journal of Biotechnology. 2010; 9(54):9255–9269.

48. Kagale S, Rozwadowski K. EAR motif-mediated transcriptional repression in plants: an underlying mechanism for epigenetic regulation of gene expression. Epigenetics, 2011; 6(2):141–146. doi: 10.4161/epi.6.2.13627 20935498

49. Liang Y, Li X, Zhang D, Gao B, Yang H, Wang Y, et al. ScDREB8, a novel A-5 type of DREB gene in the desert moss Syntrichia caninervis, confers salt tolerance to Arabidopsis. Plant physiology and biochemistry. 2017; 120:242–251. doi: 10.1016/j.plaphy.2017.09.014 29073539

50. Nakabayashi R, Yonekura-Sakakibara K, Urano K, Suzuki M, Yamada Y, Nishizawa T, et al. Enhancement of oxidative and drought tolerance in Arabidopsis by overaccumulation of antioxidant flavonoids. The Plant Journal. 2014; 77(3):367–379. doi: 10.1111/tpj.12388 24274116

51. Misyura M, Colasanti J, Rothstein S J. Physiological and genetic analysis of Arabidopsis thaliana anthocyanin biosynthesis mutants under chronic adverse environmental conditions. Journal of experimental botany. 2012; 64(1):229–240. doi: 10.1093/jxb/ers328 23162120

52. Gill SS, Tuteja N. Reactive oxygen species and antioxidant machinery in abiotic stress tolerance in crop plants. Plant physiology and biochemistry. 2010; 48(12):909–930. doi: 10.1016/j.plaphy.2010.08.016 20870416

53. Nishiyama Y, Murata N. Revised scheme for the mechanism of photoinhibition and its application to enhance the abiotic stress tolerance of the photosynthetic machinery. Applied microbiology and biotechnology. 2014; 98(21):8777–8796. doi: 10.1007/s00253-014-6020-0 25139449

54. Winkel-Shirley B. Biosynthesis of flavonoids and effects of stress. Curr Opin Plant Biol. 2002; 5:218–223. doi: 10.1016/S1369-5266(02)00256-X 11960739

55. Quan LJ, Zhang B, Shi WW, Li HY. Hydrogen peroxide in plants: a versatile molecule of the reactive oxygen species network. Journal of Integrative Plant Biology. 2008; 50(1):2–18. doi: 10.1111/j.1744-7909.2007.00599.x 18666947

Článek vyšel v časopise


2019 Číslo 10