Skip to main content
Log in

Drought-inducible expression of Hv-miR827 enhances drought tolerance in transgenic barley

  • Original Article
  • Published:
Functional & Integrative Genomics Aims and scope Submit manuscript

Abstract

Drought is one of the major abiotic stresses reducing crop yield. Since the discovery of plant microRNAs (miRNAs), considerable progress has been made in clarifying their role in plant responses to abiotic stresses, including drought. miR827 was previously reported to confer drought tolerance in transgenic Arabidopsis. We examined barley (Hordeum vulgare L. ‘Golden Promise’) plants over-expressing miR827 for plant performance under drought. Transgenic plants constitutively expressing CaMV-35S::Ath-miR827 and drought-inducible Zm-Rab17::Hv-miR827 were phenotyped by non-destructive imaging for growth and whole plant water use efficiency (WUEwp). We observed that the growth, WUEwp, time to anthesis and grain weight of transgenic barley plants expressing CaMV-35S::Ath-miR827 were negatively affected in both well-watered and drought-treated growing conditions compared with the wild-type plants. In contrast, transgenic plants over-expressing Zm-Rab17::Hv-miR827 showed improved WUEwp with no growth or reproductive timing change compared with the wild-type plants. The recovery of Zm-Rab17::Hv-miR827 over-expressing plants also improved following severe drought stress. Our results suggest that Hv-miR827 has the potential to improve the performance of barley under drought and that the choice of promoter to control the timing and specificity of miRNA expression is critical.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Subscribe and save

Springer+ Basic
EUR 32.99 /Month
  • Get 10 units per month
  • Download Article/Chapter or Ebook
  • 1 Unit = 1 Article or 1 Chapter
  • Cancel anytime
Subscribe now

Buy Now

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6

Similar content being viewed by others

References

  • Aukerman M, Park W (2009) Drought tolerant plants and related constructs and methods involving genes encoding mir827, Patent application number: US20090165168

    Google Scholar 

  • Axtell MJ, Bartel DP (2005) Antiquity of microRNAs and their targets in land plants. Plant Cell 17:1658–1673

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Bao Y, Wang C, Jiang C, Pan J, Zhang G, Liu H, Zhang H (2014) The tumor necrosis factor receptor-associated factor (TRAF)-like family protein SEVEN IN ABSENTIA 2 (SINA2) promotes drought tolerance in an ABA-dependent manner in Arabidopsis. New Phytol 202(1):174–187

  • Bartel DP (2009) MicroRNAs: target recognition and regulatory functions. Cell 136:215–233

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Blum A (2011) Drought resistance—is it really a complex trait? Funct Plant Biol 38:753–757

    Article  Google Scholar 

  • Boyer JS (1970) Leaf enlargement and metabolic rates in corn, soybean, and sunflower at various leaf water potentials. Plant Physiol 46:233–235

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Boyer JS, Westgate ME (2004) Grain yields with limited water. J Exp Bot 55:2385–2394

    Article  CAS  PubMed  Google Scholar 

  • Brien CJ (2011) dae: functions useful in the design and ANOVA of experiments. Version 2.1-7. http://cran.r-project.org/

  • Budak H, Kantar M, Bulut R, Akpinar BA (2015) Stress responsive miRNAs and isomiRs in cereals. Plant Sci 235:1–13

    Article  CAS  PubMed  Google Scholar 

  • Busk PK, Jensen AB, Pages M (1997) Regulatory elements in vivo in the promoter of the abscisic acid responsive gene rab17 from maize. Plant J 11:1285–1295

    Article  CAS  PubMed  Google Scholar 

  • Butler D, Cullis BR, Gilmour AR, Gogel BJ (2010) Analysis of mixed models for S language environments: ASReml-R reference manual. DPI Publications, Brisbane

    Google Scholar 

  • Centritto M, Lauteri M, Monteverdi MC, Serraj R (2009) Leaf gas exchange, carbon isotope discrimination, and grain yield in contrasting rice genotypes subjected to water deficits during the reproductive stage. J Exp Bot 60:2325–2339

    Article  CAS  PubMed  Google Scholar 

  • Cheah BH, Nadarajah K, Divate MD, Wickneswari R (2015) Identification of four functionally important microRNA families with contrasting differential expression profiles between drought-tolerant and susceptible rice leaf at vegetative stage. BMC Genomics 16:692–709

    Article  PubMed  PubMed Central  Google Scholar 

  • Chen C, Ridzon DA, Broomer AJ, Zhou Z, Lee DH, Nguyen JT, Barbisin M, Xu NL, Mahuvakar VR, Andersen MR, Lao KQ, Livak KJ, Guegler KJ (2005) Real-time quantification of microRNAs by stem-loop RT-PCR. Nucleic Acids Res 33:e179

    Article  PubMed  PubMed Central  Google Scholar 

  • Coombes NE (2009) Digger design search tool in R. http://www.austatgen.org/les/software/downloads/

  • Curtis MD, Grossniklaus U (2003) A gateway cloning vector set for high-throughput functional analysis of genes in planta. Plant Physiol 133:462–469

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Edwards K, Johnstone C, Thompson C (1991) A simple and rapid method for the preparation of plant genomic DNA for PCR analysis. Nucleic Acids Res 19(6):1349–1349

  • Ferdous J, Li Y, Reid N, Langridge P, Shi BJ, Tricker PJ (2015) Identification of reference genes for quantitative expression analysis of microRNAs and mRNAs in barley under various stress conditions. PLoS One 10:e0118503

    Article  PubMed  PubMed Central  Google Scholar 

  • Ferdous J, Sanchez-Ferrero JC, Langridge P, Milne L, Chowdhury J, Brien C, Tricker PJ (2016) Differential expression of microRNAs and potential targets under drought stress in barley. Plant Cell Environ. doi:10.1111/pce.12764

    PubMed  Google Scholar 

  • Frazier TP, Sun G, Burklew CE, Zhang B (2011) Salt and drought stresses induce the aberrant expression of microRNA genes in tobacco. Mol Biotechnol 49:159–165

    Article  CAS  PubMed  Google Scholar 

  • Hackenberg M, Huang P-J, Huang C-Y, Shi B-J, Gustafson P, Langridge P (2013) A comprehensive expression profile of microRNAs and other classes of non-coding small RNAs in barley under phosphorus-deficient and -sufficient consitions. DNA Res 20:109–125

    Article  CAS  PubMed  Google Scholar 

  • Hackenberg M, Gustafson P, Langridge P, Shi BJ (2015) Differential expression of microRNAs and other small RNAs in barley between water and drought conditions. Plant Biotechnol J 13:2–13

    Article  CAS  PubMed  Google Scholar 

  • Honsdorf N, March TJ, Berger B, Tester M, Pillen K (2014) High-throughput phenotyping to detect drought tolerance qtl in wild barley introgression lines. PLoS One 9:e97047

    Article  PubMed  PubMed Central  Google Scholar 

  • Ismagul A, Mazonka I, Callegari C, Eliby S (2014) Agrobacterium-mediated transformation of barley (Hordeum vulgare L.). In: Fleury D, Whitford R (eds) Crop breeding methods and protocols, 1st edn. Springer, New York, pp 203–211

    Chapter  Google Scholar 

  • Jefferson RA, Kavanagh TA, Bevan MW (1987) GUS fusion: β-glucuronidase as a sensitive and versatile gene fusion marker in higher plants. EMBO J 6:3901–3907

    CAS  PubMed  PubMed Central  Google Scholar 

  • Jones-Rhoades MW, Bartel DP, Bartel B (2006) MicroRNAs and their regulatory roles in plants. Annu Rev Plant Biol 57:19–53

    Article  CAS  PubMed  Google Scholar 

  • Kantar M, Lucas SJ , Budak H, Kantar M, Lucas SJ, Budak H (2011) miRNA expression patterns of Triticum dicoccoides in response to shock drought stress. Planta 233(3):471–484

  • Kasuga M, Liu Q, Miura S, Yamaguchi-Shinozaki K, Shinozaki K (1999) Improving plant drought, salt, and freezing tolerance by gene transfer of a single stress-inducible transcription factor. Nat Biotechnol 17:287–291

    Article  CAS  PubMed  Google Scholar 

  • Katiyar A, Smita S, Muthusamy SK, Chinnusamy V, Pandey DM, Bansal KC (2015) Identification of novel drought-responsive microRNAs and trans-acting siRNAs from Sorghum bicolor (L.) Moench by high-throughput sequencing analysis. Front Plant Sci 6:1–21

    Article  Google Scholar 

  • Kulcheski FR, de Oliveira LFV, Molina LG, Almerão MP, Rodrigues FA, Marcolino J, Barbosa JF, Stolf-Moreira R, Nepomuceno AL, Marcelino-Guimarães FC, Abdelnoor RV, Nascimento LC, Carazzolle MF, Pereira GAG, Margis R (2011) Identification of novel soybean microRNAs involved in abiotic and biotic stresses. BMC Genomics 12:307–323

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Lawlor DW (2013) Genetic engineering to improve plant performance under drought: physiological evaluation of achievements, limitations, and possibilities. J Exp Bot 64:83–108

    Article  CAS  PubMed  Google Scholar 

  • Li WX, Oono Y, Zhu J, He XJ, Wu JM, Iida K, Lu XY, Cui X, Jin H, Zhu JK (2008) The Arabidopsis NFYA5 transcription factor is regulated transcriptionally and posttranscriptionally to promote drought resistance. Plant Cell 20:2238–2251

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Lin SI, Santi C, Jobet E, Lacut E, El Kholti N, Karlowski WM, Verdeil JL, Breitler JC, Périn C, Ko SS, Guiderdoni E, Chiou TJ, Echeverria M (2010) Complex regulation of two target genes encoding SPX-MFS proteins by rice miR827 in response to phosphate starvation. Plant Cell Physiol 51:2119–2131

    Article  CAS  PubMed  Google Scholar 

  • Liu HH, Tian X, Li YJ, Wu CA, Zheng CC (2008) Microarray-based analysis of stress-regulated microRNAs in Arabidopsis thaliana. RNA 14:836–843

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Liu Q, Wang F, Axtell MJ (2014) Analysis of complementarity requirements for plant microRNA targeting using a Nicotiana benthamiana quantitative transient assay. Plant Cell 26:741–753

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Liu H, Searle IR, Watson-Haigh NS, Baumann U, Mather DE, Able AJ, Able JA (2015) Genome-wide identification of microRNAs in leaves and the developing head of four durum genotypes during water deficit stress. PLoS One. doi:10.1371/journal.pone.0142799

    Article  Google Scholar 

  • Liu H, Able AJ, Able JA (2016a) Water-deficit responsive microRNAs and their targets in four durum wheat genotypes. Funct Integr Genomics. doi:10.1007/s10142-016-0515-y

    Google Scholar 

  • Liu H, Able AJ, Able JA (2016b) SMARTER de-stressed cereal breeding. Trends Plant Sci. doi:10.1016/j.tplants.2016.07.006

    PubMed  Google Scholar 

  • Lu XY, Huang XL (2008) Plant miRNAs and abiotic stress responses. Biochem Biophys Res Commun 368:458–462

    Article  CAS  PubMed  Google Scholar 

  • Morran S, Eini O, Pyvovarenko T, Parent B, Singh R, Ismagul A, Eliby S, Shirley N, Langridge P, Lopato S (2011) Improvement of stress tolerance of wheat and barley by modulation of expression of DREB/CBF factors. Plant Biotechnol J 9:230–249

    Article  CAS  PubMed  Google Scholar 

  • Nakashima K, Ito Y, Yamaguchi-Shinozaki K (2009) Transcriptional regulatory networks in response to abiotic stresses in Arabidopsis and grasses. Plant Physiol 149:88–95

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Nevo E, Chen G (2010) Drought and salt tolerances in wild relatives for wheat and barley improvement. Plant Cell Environ 33:670–685

    Article  CAS  PubMed  Google Scholar 

  • Oh SJ, Kwon CW, Choi DW, Song SI, Kim JK (2007) Expression of barley HvCBF4 enhances tolerance to abiotic stress in transgenic rice. Plant Biotechnol J 5:646–656

    Article  CAS  PubMed  Google Scholar 

  • Parizotto EA, Dunoyer P, Rahm N, Himber C, Voinnet O (2004) In vivo investigation of the transcription, processing, endonucleolytic activity, and functional relevance of the spatial distribution of a plant miRNA. Genes Dev 18:2237–2242

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Qin Y, Duan Z, Xia X, Yin W (2011) Expression profiles of precursor and mature microRNAs under dehydration and high salinity shock in Populus euphratica. Plant Cell Rep 30:1893–1907

    Article  CAS  PubMed  Google Scholar 

  • R Development Core Team (2014) R: a language and environment for statistical computing. R Foundation for Statistical Computing, Vienna, http://www.r-project.org/

    Google Scholar 

  • R Development Core Team (2015) R: a language and environment for statistical computing. R Foundation for Statistical Computing, Vienna, http://www.r-project.org/

    Google Scholar 

  • Schreiber AW, Shi B-J, Huang C-Y, Langridge P, Baumann U (2011) Discovery of barley miRNAs through deep sequencing of short reads. BMC Genomics 12:129

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Schwab R, Palatnik JF, Riester M, Schommer C, Schmid M, Weigel D (2005) Specific effects of microRNAs on the plant transcriptome. Dev Cell 8:517–527

    Article  CAS  PubMed  Google Scholar 

  • Shi BJ, Sutton T, Collins NC, Pallotta M, Langridge P (2010) Construction of a barley bacterial artificial chromosome library suitable for cloning genes for boron tolerance, sodium exclusion and high grain zinc content. Plant Breed 129:291–296

    Article  CAS  Google Scholar 

  • Shinozaki K, Yamaguchi-Shinozaki K (2007) Gene networks involved in drought stress response and tolerance. J Exp Bot 58:221–227

    Article  CAS  PubMed  Google Scholar 

  • Sunkar R, Zhu JK (2004) Novel and stress-regulated microRNAs and other small RNAs from Arabidopsis. Plant Cell 16:2001–2019

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Sunkar R, Chinnusamy V, Zhu J, Zhu JK (2007) Small RNAs as big players in plant abiotic stress responses and nutrient deprivation. Trends Plant Sci 12:301–309

    Article  CAS  PubMed  Google Scholar 

  • Sunkar R, Li YF, Jagadeeswaran G (2012) Functions of microRNAs in plant stress responses. Trends Plant Sci 17:196–203

    Article  CAS  PubMed  Google Scholar 

  • Trindade I, Capitao C, Dalmay T, Fevereiro MP, dos Santos DM (2010) miR398 and miR408 are up-regulated in response to water deficit in Medicago truncatula. Planta 231:705–716

    Article  CAS  PubMed  Google Scholar 

  • Varkonyi-Gasic E, Wu R, Wood M, Walton EF, Hellens RP (2007) Protocol: a highly sensitive RT-PCR method for detection and quantification of microRNAs. Plant Methods 3:12–23

    Article  PubMed  PubMed Central  Google Scholar 

  • Wei L, Zhang D, Xiang F, Zhang Z (2009) Differentially expressed miRNAs potentially involved in the regulation of defense mechanism to drought stress in maize seedlings. Int J Plant Sci 170:979–989

    Article  CAS  Google Scholar 

  • Xu Z, Dooner HK (2006) The maize aberrant pollen transmission 1 gene is a SABRE/KIP homolog required for pollen tube growth. Genetics 172:1251–1261

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Xu Z, Zhou G, Shimizu H (2010) Plant responses to drought and rewatering. Plant Signaling Behav 5:649–654

    Article  CAS  Google Scholar 

  • Zhang JZ, Creelman RA, Zhu JK (2004) From laboratory to field: using information from Arabidopsis to engineer salt, cold, and drought tolerance in crops. Plant Physiol 135:615–621

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Zhang B, Pan X, Cannon CH, Cobb GP, Anderson TA (2006) Conservation and divergence of plant microRNA genes. Plant J 46:243–259

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgments

This research was supported by a grant to the Australian Centre for Plant Functional Genomics, supported through research funding from DuPont/Pioneer (USA). Our grateful thanks to Patricia Warner and ACPFG Transformation Group for barley transformation; Margaret Pallotta and Suzanne Manning for their assistance with Southern blotting; and Dr. Sergiy Lopato, Dr. Ainur Ismagul and Dr. Nataliya Kovalchuk for generation and selection of the Zm-Rab17::GUS transgenic barley germplasm. We specially thank the team of The Plant Accelerator for technical support in running the experiment and conducting the image analysis. The Plant Accelerator, Australian Plant Phenomics Facility, is funded under the National Collaborative Infrastructure Strategy. We are thankful to Dr. Ursula Langridge, Alex Kovalchuk and Yuri Onyskiv for their assistance with growing plants at different phases of this experiment, and Yuan Li and Hui Zhou for performing the qRT-PCR assays. We also thank Dr. Bu-Jun Shi for his advice at the early stage of this experiment.

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Penny J. Tricker.

Additional information

This article forms part of a special issue of Functional & Integrative Genomics entitled “miRNA in model and complex organisms” (Issue Editors: Hikmet Budak and Baohong Zhang)

Electronic supplementary materials

Below is the link to the electronic supplementary material.

ESM 1

(XLSX 20 kb)

ESM 2

(DOCX 23365 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Ferdous, J., Whitford, R., Nguyen, M. et al. Drought-inducible expression of Hv-miR827 enhances drought tolerance in transgenic barley. Funct Integr Genomics 17, 279–292 (2017). https://doi.org/10.1007/s10142-016-0526-8

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s10142-016-0526-8

Keywords

Navigation