Skip to main content
SpringerLink
Log in

Functional Analysis of Two Orthologous NAC Genes, CarNAC3, and CarNAC6 from Cicer arietinum, Involved in Abiotic Stresses in Poplar

  • Original Paper
  • Published:
Plant Molecular Biology Reporter Aims and scope Submit manuscript

Abstract

Abiotic stresses such as drought and high salinity inhibit plant growth and productivity. NAC transcription factors play a variety of important roles in plant development and abiotic stress responses. To date, the transformation of NAC genes into poplar plants has not been reported. In this study, we introduced the CarNAC3 and CarNAC6 salinity and drought tolerant genes from Cicer arietinum (chickpea) into Populus deltoides × Populus euramericana ‘Nanlin895’ (poplar) plants using Agrobacterium tumefaciens-mediated transformation. We verified the integration of the two genes into the poplar genome using polymerase chain reaction (PCR) and their stable expression was confirmed at the transcript level using real-time (RT)-PCR. The growth rates of roots and shoots and the clawed root rate increased in transformed plants grown in agar medium or in soil and increased in response to drought and salt stress conditions. Proline and photoprotectant pigment accumulation and antioxidant enzyme activities increased in response to abiotic stress, but the accumulation of photosynthetic pigments and malondialdehyde (MDA) decreased in transgenic lines relative to wild-type control plants. The CarNAC3 transgene was expressed at higher levels than the CarNAC6 transgene. Our results showed that the CarNAC3 and CarNAC6 genes enhanced the capacity for osmotic adjustment and increased antioxidant enzyme activity and suggest that these genes could play a significant role in improving drought and salt tolerance when expressed in poplar plants.

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

Access this article

Log in via an institution

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

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7

Similar content being viewed by others

Abbreviations

6-BA:

N-6-benzyladenine

TDZ:

Thidiazuron

AS:

Acetosyringone

NPTII :

Neomycin phosphotransferase

MS:

Murashige and Skoog

ORF:

Open reading frame

PCR:

Polymerase chain reaction

RT-PCR:

Reverse transcription polymerase chain reaction

MDA:

Malondialdehyde

POD:

Guaiacol peroxidase

SOD:

Superoxide dismutase

ROS:

Reactive oxygen species

References

  • Abdullahil BM, Lee EJ, Paek KY (2010) Medium salt strength induced changes in growth, physiology and secondary metabolite content in adventitious roots of Morinda citrifolia: the role of antioxidant enzymes and phenylalanine ammonia lyase. Plant Cell Rep 29:685–694

    Article  Google Scholar 

  • Abraham E, Hourton-Cabassa C, Erdei L, Szabados L (2010) Methods for determination of proline in plants. Methods Mol Biol 639:317–331

    Article  CAS  PubMed  Google Scholar 

  • Ahmad MSA, Ashraf M, Ali Q (2010) Soil salinity as a selection pressure is a key determinant for the evolution of salt tolerance in blue panic grass (Panicum antidotale Retz.). Flora 205:37–45

    Article  Google Scholar 

  • Aida M, Ishida T, Fukaki H, Fujisawa H, Tasaka M (1997) Genes involved in organ separation in Arabidopsis: an analysis of the cup-shaped cotyledon mutant. Plant Cell 9:841–857

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Al-Whaibi MH, Siddiqui MH, Basalah MO (2011) Salicylic acid and calcium-induced protection of wheat against salinity. Protoplasma 249:769–778

    Article  PubMed  Google Scholar 

  • Barba-Espin G, Clemente-Moreno MJ, Alvarez S, Garcia-Legaz MF, Hernandez JA, Diaz-Vivancos P (2011) Salicylic acid negatively affects the response to salt stress in pea plants. Plant Biol (Stuttg) 13:909–917

    Article  CAS  Google Scholar 

  • Ben AC, Ben RB, Sensoy S, Boukhriss M, Ben AF (2010) Exogenous proline effects on photosynthetic performance and antioxidant defense system of young olive tree. J Agric Food Chem 58:4216–4222

    Article  Google Scholar 

  • Bhagat SS, Ghone RA, Suryakar AN, Hundekar PS (2011) Lipid peroxidation and antioxidant vitamin status in colorectal cancer patients. Indian J Physiol Pharmacol 55:72–76

    CAS  PubMed  Google Scholar 

  • Bursy J, Pierik AJ, Pica N, Bremer E (2007) Osmotically induced synthesis of the compatible solute hydroxyectoine is mediated by an evolutionarily conserved ectoine hydroxylase. J Biol Chem 282:31147–31155

    Article  CAS  PubMed  Google Scholar 

  • Cho K, Kim YC, Woo JC, Rakwal R, Agrawal GK, Yoeun S, Han O (2012) Transgenic expression of dual positional maize lipoxygenase-1 leads to the regulation of defense-related signaling molecules and activation of the antioxidative enzyme system in rice. Plant Sci 185–186:238–245

    Article  PubMed  Google Scholar 

  • Collinge M, Boller T (2001) Differential induction of two potato genes, Stprx2 and StNAC, in response to infection by Phytophthora infestans and to wounding. Plant Mol Biol 46:521–529

    Article  CAS  PubMed  Google Scholar 

  • Dong J, Wan G, Liang Z (2010) Accumulation of salicylic acid-induced phenolic compounds and raised activities of secondary metabolic and antioxidative enzymes in Salvia miltiorrhiza cell culture. J Biotechnol 148:99–104

    Article  CAS  PubMed  Google Scholar 

  • Fowler S, Thomashow MF (2002) Arabidopsis transcriptome profiling indicates that multiple regulatory pathways are activated during cold acclimation in addition to the CBF cold response pathway. Plant Cell 14:1675–1690

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Guo YF, Gan SS (2006) AtNAP, a NAC family transcription factor, has an important role in leaf senescence. Plant J 46:601–612

    Article  CAS  PubMed  Google Scholar 

  • He XJ, Mu RL, Cao WH, Zhang ZG, Zhang JS, Chen SY (2005) AtNAC2, a transcription factor downstream of ethylene and auxin signaling pathways, is involved in salt stress response and lateral root development. Plant J 44:903–916

    Article  CAS  PubMed  Google Scholar 

  • Hegedus D, Yu M, Baldwin D, Gruber M, Sharpe A, Parkin I, Whitwill S, Lydiate D (2003) Molecular characterization of Brassica napus NAC domain transcriptional activators induced in response to biotic and abiotic stress. Plant Mol Biol 53:383–397

    Article  CAS  PubMed  Google Scholar 

  • Hoekema A, Hirsch PR, Hooykaas PJJ, Schilperoort RA (1983) A binary plant vector strategy based on separation of vir- and T-region of the Agrobacterium tumefaciens Ti plasmid. Nature 303:179–180

    Article  CAS  Google Scholar 

  • Horsch RB, Fry J, Hoffmann N, Neidermeyer J, Rogers SG, Fraley RT (1988) Leaf disc transformation: plant molecular biology manual. Kluwer Academic Publishers, Dordrecht, pp 1–9

    Google Scholar 

  • Hou XN, Liang YZ, He XL, Shen YZ, Huang ZJ (2013) A novel ABA responsive TaSRHP gene from wheat contributes to enhanced resistance to salt stress in Arabidopsis thaliana. Plant Mol Biol Rep 31:791–801

    Article  CAS  Google Scholar 

  • Hu H, Dai M, Yao J, Xiao B, Li X, Zhang Q, Xiong L (2006) Overexpressing a NAM, ATAF, and CUC (NAC) transcription factor enhances drought resistance and salt tolerance in rice. Proc Natl Acad Sci U S A 103:12987–12992

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Hui P, Cheng HY, Chen C, Yu XW, Yang JN, Gao WR, Shi QH, Zhang H, Li JG, Ma H (2009) A NAC transcription factor gene of chickpea (Cicer arietinum), CarNAC3, is involved in drought stress response and various developmental processes. J Plant Physiol 166:1934–1945

    Article  Google Scholar 

  • Krantev A, Yordanova R, Janda T, Szalai G, Popova L (2008) Treatment with salicylic acid decreases the effect of cadmium on photosynthesis in maize plants. J Plant Physiol 165:920–931

    Article  CAS  PubMed  Google Scholar 

  • Lei Y, Korpelainen H, Li C (2007) Physiological and biochemical responses to high Mn concentrations in two contrasting Populus cathayana populations. Chemosphere 68:686–694

    Article  CAS  PubMed  Google Scholar 

  • Lichtenthaler HK (1987) Chlorophylls and carotenoids: pigments of photosynthetic biomembranes. Methods Enzymol 148:350–382

    Article  CAS  Google Scholar 

  • Logemann E, Birkenbih RP, Ülker B, Somssich IE (2006) An improved method for preparing Agrobacterium cells that simplifies the Arabidopsis transformation protocol. Plant Methods 2:16. doi:10.1186/1746-4811-2-16

    Article  PubMed Central  PubMed  Google Scholar 

  • Meng QC, Zhang CH, Gai JY, Yu DY (2007) Molecular cloning, sequence characterization and tissue-specific expression of six NAC-like genes in soybean (Glycine max (L.) Merr.). J Plant Physiol 51:617–30

    Google Scholar 

  • Mishra P, Bhoomika K, Dubey RS (2011) Differential responses of antioxidative defense system to prolonged salinity stress in salt tolerant and salt-sensitive Indica rice (Oryza sativa L.) seedlings. Protoplasma. doi:10.1007/s00709-011-0365-3

    Google Scholar 

  • Mitsuda N, Iwase A, Yamamoto H, Yoshida M, Seki M, Shinozaki K, Ohme-Takagi M (2007) NAC transcription factors, NST1 and NST3, are key regulators of the formation of secondary walls in woody tissues of Arabidopsis. Plant Cell 19:270–280

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Mittova V, Guy M, Tal M, Volokita M (2004) Salinity up-regulates the antioxidative system in root mitochondria and peroxisomes of the wild salt-tolerant tomato species Lycopersicon pennellii. J Exp Bot 55:1105–1113

    Article  CAS  PubMed  Google Scholar 

  • Movahedi A, Zhang J, Gao P, Yang Y, Wang L, Yin T, Kadkhodaei S, Ebrahimi M, Zhuge Q (2014a) Expression of the chickpea CarNAC3 gene enhances salinity and drought tolerance in transgenic poplars. Plant Cell Tiss Organ Cult. doi:10.1007/s11240-014-0588-z

    Google Scholar 

  • Movahedi A, Zhang J, Amirian R, Zhuge Q (2014b) An efficient Agrobacterium-mediated transformation system for poplar. Int J Mol Sci 15(6):10780–10793

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Mysore KS, Crasta OR, Tuori RP, Folkerts O, Swirsky PB, Martin GB (2002) Comprehensive transcript profiling of Pto- and Prf-mediated host defense responses to infection by Pseudomonas syringae pv. tomato. Plant J 32:299–315

    Article  CAS  PubMed  Google Scholar 

  • Nakashima K, Tran LS, Van Nguyen D, Fujita M, Maruyama K, Todaka D, Ito Y, Hayashi N, Shinozaki K, Yamaguchi-Shinozaki K (2007) Functional analysis of a NAC type transcription factor OsNAC6 involved in abiotic and biotic stress responsive gene expression in rice. Plant J 51:617–630

    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  PubMed Central  CAS  PubMed  Google Scholar 

  • Nakashima K, Takasaki H, Mizoi J, Shinozaki K, Yamaguchi-Shinozaki K (2012) NAC transcription factors in plant abiotic stress responses. Biochimica et Biophysica Acta (BBA) - Gene Regulatory Mechanisms 1819:97–103

    Article  CAS  Google Scholar 

  • Nounjan N, Nghia PT, Theerakulpisut P (2012) Exogenous proline and trehalose promote recovery of rice seedlings from salt-stress and differentially modulate antioxidant enzymes and expression of related genes. J Plant Physiol 169:596–604

    Article  CAS  PubMed  Google Scholar 

  • Olsen AN, Ernst HA, Leggio LL, Skriver K (2005) NAC transcription factors: structurally distinct, functionally diverse. Trends Plant Sci 10:79–87

    Article  CAS  PubMed  Google Scholar 

  • Ooka H, Satoh K, Doi K, Nagata T, Otomo Y, Murakami K et al (2003) Comprehensive analysis of NAC family genes in Oryza sativa and Arabidopsis thaliana. DNA Res 10:239–47

    Article  CAS  PubMed  Google Scholar 

  • Pagariya MC, Devarumath RM, Kawar PG (2012) Biochemical characterization and identification of differentially expressed candidate genes in salt-stressed sugarcane. Plant Sci 184:1–13

    Article  CAS  PubMed  Google Scholar 

  • Porebski L, Bailey L, Baum B (1997) Modification of a CTAB DNA extraction protocol for plants containing high polysaccharide and polyphenol components. Plant Mol Biol Rep 15:8–15

    Article  CAS  Google Scholar 

  • Rabbani MA, Maruyama K, Abe H, Khan MA, Katsura K, Ito Y, Yoshiwara K, Seki M, Shinozaki K, Yamaguchi-Shinozaki K (2003) Monitoring expression profiles of rice genes under cold, drought, and high-salinity stresses and abscisic acid application using cDNA microarray and RNA gel-blot analyses. Plant Physiol 133:1755–1767

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Sablowski RW, Meyerowitz EM (1998) A homolog of No Apical Meristem is an immediate target of the floral homeotic genes APETALA3/PISTILLATA. Cell 92:93–103

    Article  CAS  PubMed  Google Scholar 

  • Santner A, Estelle M (2009) Recent advances and emerging trends in plant hormone signalling. Nature 459:1071–1078

    Article  CAS  PubMed  Google Scholar 

  • Satoh K (1978) Serum lipid peroxide in cerebrovascular disorders determined by a new colorimetric method. Clin Chim Acta 90:37–43

    Article  CAS  PubMed  Google Scholar 

  • Song S, Liu W, Guo S, Shang Q, Zhang Z (2006) Salt resistance and its mechanism of cucumber under effects of exogenous chemical activators. Ying Yong Sheng Tai Xue Bao 17:1871–1876

    CAS  PubMed  Google Scholar 

  • Souer E, Van AH, Kloos D, Mol J, Koes R (1996) The No Apical Meristem gene of petunia is required for pattern formation in embryos and flowers and is expressed at meristem and primordia boundaries. Cell 85:159–170

    Article  CAS  PubMed  Google Scholar 

  • Stewart RR, Bewley JD (1980) Lipid peroxidation associated with accelerated aging of soybean axes. Plant Physiol 65:245–248

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Su X, Chu Y, Li H, Hou Y, Zhang B, Huang Q, Hu Z, Huang R, Tian Y (2011) Expression of multiple resistance genes enhances tolerance to environmental stressors in transgenic poplar (Populus x euramericana ‘Guariento’). PLoS ONE 6:e24614

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Takasaki H, Maruyama K, Kidokoro S, Ito Y, Fujita Y, Shinozaki K, Yamaguchi-Shinozaki K, Nakashima K (2010) The abiotic stress-responsive NAC-type transcription factor OsNAC5 regulates stress-inducible genes and stress tolerance in rice. Mol Genet Genomics 284:173–183

    Article  CAS  PubMed  Google Scholar 

  • Uauy C, Distelfeld A, Fahima T, Blechl A, Dubcovsky J (2006) A NAC gene regulating senescence improves grain protein, zinc, and iron content in wheat. Science 314:1298–301

    Article  CAS  PubMed  Google Scholar 

  • Wang N, Hua H, Egrinya Eneji A, Li Z, Duan L, Tian X (2012) Genotypic variations in photosynthetic and physiological adjustment to potassium deficiency in cotton (Gossypium hirsutum). J Photochem Photobiol B 110:1–8

    Article  CAS  PubMed  Google Scholar 

  • Xie Q, Sanz-Burgos AP, Guo H, García JA, Gutiérrez C (1999) GRAB proteins, novel members of the NAC domain family, isolated by their interaction with a Gemini virus protein. Plant Mol Biol 39:647–656

    Article  CAS  PubMed  Google Scholar 

  • Yamaguchi-Shinozaki K, Shinozaki K (2006) Transcriptional regulatory networks in cellular responses and tolerance to dehydration and cold stresses. Annu Rev Plant Biol 57:781–803

    Article  CAS  PubMed  Google Scholar 

  • Yamaguchi-Shinozaki K, Koizumi M, Urao S, Shinozaki K (1992) Molecular cloning and characterization of 9 cDNAs for genes that are responsive to desiccation in Arabidopsis thaliana: sequence analysis of one cDNA clone that encodes a putative transmembrane channel protein. Plant Cell Physiol 33:217–224

    CAS  Google Scholar 

  • Zheng XN, Chen B, Lu GJ, Han B (2009) Overexpression of a NAC transcription factor enhances rice drought and salt tolerance. Biochem Biophys Res Commun 379:985–989

    Article  CAS  PubMed  Google Scholar 

  • Zhou J, Wang J, Bi Y, Wang L, Tang L, Yu X, Ohtani M, Demura T, Zhuge Q (2014) Overexpression of PtSOS2 enhances salt tolerance in transgenic poplars. Plant Mol Biol Rep 32:185–197

    Article  PubMed Central  CAS  PubMed  Google Scholar 

Download references

Acknowledgments

We thank Prof. Hao Ma (Nanjing Agricultural University, China) for providing the CarNAC3 and CarNAC6 plasmids. This work was supported by grants from the Forestry Public Benefic Research Program (No. 201304102) and the International Science and Technology Cooperation Program of China (No. 2014DFG32440), Priority Academic Program Development of Jiangsu Higher Education Institution, and the Program for Innovative Research Team at the University of Educational Department and Jiangsu Province, China.

Conflict of Interest

The authors do not have conflicts of interest to declare.

Author information

Authors and Affiliations

  1. Co-Innovation Center for Sustainable Forestry in Southern China, Nanjing Forestry University, Nanjing, 210037, China

    Ali Movahedi, Jiaxin Zhang, Tongming Yin & Qiang Zhuge

Authors
  1. Ali Movahedi
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Jiaxin Zhang
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Tongming Yin
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Qiang Zhuge
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Qiang Zhuge.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Supplemental Data 1

(DOC 681 kb)

Supplemental Data 2

(DOC 69 kb)

Supplemental Data 3

(DOC 906 kb)

Supplemental Data 4

(DOC 589 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Movahedi, A., Zhang, J., Yin, T. et al. Functional Analysis of Two Orthologous NAC Genes, CarNAC3, and CarNAC6 from Cicer arietinum, Involved in Abiotic Stresses in Poplar. Plant Mol Biol Rep 33, 1539–1551 (2015). https://doi.org/10.1007/s11105-015-0855-0

Download citation

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s11105-015-0855-0

Keywords

Search

Navigation