Advertisement

Arabidopsis thaliana DHS2 (AT4G33510) gene promoter is highly wound responsive and requires a part of the first exon sequences for its function

  • Ritesh Kumar Raipuria
  • Vajinder Kumar
  • Kadur Narayan Guruprasad
  • Shripad Ramachandra Bhat
Short Communication

Abstract

Wound-responsive promoters are critical for developing the next generation of pest-tolerant transgenics, which would defend host by producing defensive factors at the site of attack. As shikimate pathway genes are known to be induced upon wounding, we evaluated the promoter of Arabidopsis thaliana DHS2 gene, which encodes for the first enzyme of this pathway. Transcription start site mapping showed DHS2 transcript has 89 nucleotide long 5′ UTR and RT-PCR revealed constitutive expression of DHS2 gene. Promoter analysis in transgenic A. thaliana using uidA reporter gene revealed that a part of the first exon of DHS2 is essential for promoter function. Histochemical GUS expression studies showed DHS2 expression in all green parts with the least expression in seedling roots and pollens. DHS2 promoter was found to be highly responsive to wounding giving 2–4 log fold up regulation within 10 min of wounding. Elevated expression level remained stable for 2–3 h and returned to normal after 24 h. In silico search identified wound responsive cis elements within the first exon of the DHS2 gene further underscoring the role of coding sequences in wound response and promoter activity. This is the first report demonstrating that coding sequence of a gene is an essential part of its promoter.

Keywords

3-Deoxy-d-arabino-heptulosonate 7-phosphate synthase 2 Shikimate pathway Wound responsive promoter 

Abbreviations

DHS2

3-Deoxy-d-arabino-heptulosonate 7-phosphate synthase 2

IGR

Intergenic region

PP2C

Protein phosphatase 2C

TSS

Transcription start site

Notes

Acknowledgements

We thank NFBSRA, Indian Council of Agricultural Research, New Delhi for financial assistance and National Phytotron Facility, IARI, New Delhi for help in raising Arabidopsis plants under contained conditions.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Supplementary material

13562_2017_430_MOESM1_ESM.docx (133 kb)
Supplementary material 1 (DOCX 133 kb)

References

  1. Chen C, Chen Z (2002) Potentiation of developmentally regulated plant defense response by AtWRKY18, a pathogen-induced Arabidopsis transcription factor. Plant Physiol 129:706–716CrossRefPubMedPubMedCentralGoogle Scholar
  2. Chen Y, Zhang X, Wu W, Chen Z, Gu H, Qu LJ (2006) Overexpression of the wounding-responsive gene AtMYB15 activates the shikimate pathway in Arabidopsis. J Integr Plant Biol 48:1084–1095CrossRefGoogle Scholar
  3. Coutu C, Brandle J, Brown D, Brown K, Miki B, Simmonds J, Hegedus DD (2007) pORE, a modular binary vector series suited for both monocot and dicot plant transformation. Transgenic Res 16:771–781CrossRefPubMedGoogle Scholar
  4. Daraselia ND, Tarchevskaya S, Narita JO (1996) The promoter for tomato 3-hydroxy-3-methylglutaryl coenzyme A reductase gene 2 has unusual regulatory elements that direct high-level expression. Plant Physiol 112:727–733CrossRefPubMedPubMedCentralGoogle Scholar
  5. Dyer WE, Henstrand JM, Handa AK, Herrmann KM (1989) Wounding induces the first enzyme of the shikimate pathway in Solanaceae. Proc Natl Acad Sci 86:7370–7373CrossRefPubMedPubMedCentralGoogle Scholar
  6. Emami S, Arumainayagam D, Korf I, Rose AB (2013) The effects of a stimulating intron on the expression of heterologous genes in Arabidopsis thaliana. Plant Biotechnol J 11:555–563CrossRefPubMedGoogle Scholar
  7. Emanuelsson O, Nielsen H, Heijne GV (1999) ChloroP, a neural network-based method for predicting chloroplast transit peptides and their cleavage sites. Protein Sci 8:978–984CrossRefPubMedPubMedCentralGoogle Scholar
  8. Gallegos JE, Rose AB (2017) Intron DNA sequences can be more important than the proximal promoter in determining the site of transcript initiation. Plant Cell. doi: 10.1105/tpc.17.00020 PubMedPubMedCentralGoogle Scholar
  9. Gupta NC, Jain PK, Bhat SR, Srinivasan R (2012) Upstream sequence of fatty acyl-CoA reductase (FAR6) of Arabidopsis thaliana drives wound-inducible and stem-specific expression. Plant Cell Rep 31:839–850CrossRefPubMedGoogle Scholar
  10. Higo K, Ugawa Y, Iwamoto M, Korenaga T (1999) Plant cis-acting regulatory DNA elements (PLACE) database. Nucleic Acids Res 27:297–300CrossRefPubMedPubMedCentralGoogle Scholar
  11. Keith B, Dong X, Ausubel F, Fink G (1991) Differential induction of 3-deoxy-d-arabino-heptulosonate 7-phosphate synthase genes in Arabidopsis thaliana by wounding and pathogenic attack. Proc Natl Acad Sci USA 88:8821–8825CrossRefPubMedPubMedCentralGoogle Scholar
  12. Kumar V, Thakare DR, Saha DN, Jajoo A, Jain PK, Bhat SR, Srinivasan R (2012) Characterization of upstream sequences of the peroxidase gene, Atprx18 of Arabidopsis thaliana. J Plant Biochem Biotechnol 21:121–127CrossRefGoogle Scholar
  13. Lescot M, Déhais P, Thijs G, Marchal K, Moreau Y, Van de Peer Y, Rouzé P, Rombauts S (2002) Plant CARE, a database of plant cis-acting regulatory elements and a portal to tools for in silico analysis of promoter sequences. Nucleic Acids Res 301:325–327CrossRefGoogle Scholar
  14. Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real–time quantitative PCR and the 2−ΔΔCT method. Methods 25:402–408CrossRefPubMedGoogle Scholar
  15. Maeda H, Dudareva N (2012) The Shikimate pathway and aromatic amino acid biosynthesis in plants. Annu Rev Plant Biol 63:73–105CrossRefPubMedGoogle Scholar
  16. Nishiuchi T, Shinshi H, Suzuki K (2004) Rapid and transient activation of transcription of the ERF3 gene by wounding in tobacco leaves. J Biol Chem 279:55355–55361CrossRefPubMedGoogle Scholar
  17. Park SH, Lim H, Hyun SJ, Yun DW, Yoon UH, Ji H, Kim TH, Eun MY, Kim YH, Lee GS (2014) Wound-inducible expression of the OsDof1 gene promoter in a Ds insertion mutant and transgenic plants. Plant Biotechnol Rep 8:305–313CrossRefGoogle Scholar
  18. Parra G, Bradnam K, Rose A, Korf I (2011) Comparative and functional analysis of intron-mediated enhancement signals reveals conserved features among plants. Nucleic Acids Res 39:5328–5337CrossRefPubMedPubMedCentralGoogle Scholar
  19. Reyes JC, Muro-Pastor MI, Florencio FJ (2004) The GATA family of transcription factors in Arabidopsis and rice. Plant Physiol 134:1718–1732CrossRefPubMedPubMedCentralGoogle Scholar
  20. Rose A, Elfersi T, Parra G, Korf I (2008) Promoter-proximal introns in Arabidopsis thaliana are enriched in dispersed signals that enhance gene expression. Plant Cell 20:543–551CrossRefPubMedPubMedCentralGoogle Scholar
  21. Rouster J, Leah R, Mundy J, Cameron-Mills V (1997) Identification of a methyl jasmonate-responsive region in the promoter of a lipoxygenase 1 gene expressed in barley grain. Plant J 11:513–523CrossRefPubMedGoogle Scholar
  22. Sasaki K, Hiraga S, Ito H, Seo S, Matsui H, Ohashi Y (2002) A wound-inducible Tobacoo peroxidase gene expresses preferentially in the vascular system. Plant Cell Physiol 43:108–117CrossRefPubMedGoogle Scholar
  23. Trtikova M, Wikmark OG, Zemp N, Widmer A, Hilbeck A (2015) Transgene expression and Bt protein content in transgenic bt maize (MON810) under optimal and stressful environmental conditions. PLoS ONE 10:e0123011CrossRefPubMedPubMedCentralGoogle Scholar
  24. Tzin V, Galili G (2010) New insights into the Shikimate and aromatic amino acids biosynthesis pathways in plants. Mol Plant 3:956–972CrossRefPubMedGoogle Scholar
  25. Vijayan J, Devanna BN, Singh NK, Sharma TR (2015) Cloning and functional validation of early inducible Magnaporthe oryzae responsive CYP76M7 promoter from rice. Front Plant Sci 6:371CrossRefPubMedPubMedCentralGoogle Scholar
  26. Winter D, Vinegar B, Nahal H, Ammar R, Wilson GV, Provart NJ (2007) An “Electronic Fluorescent Pictograph” browser for exploring and analyzing large-scale biological data sets. PLoS ONE 2:e718CrossRefPubMedPubMedCentralGoogle Scholar
  27. Yanagisawa S, Schmidt RJ (1999) Diversity and similarity among recognition sequences of Dof transcription factors. Plant J 17:209–214CrossRefPubMedGoogle Scholar
  28. Zybailov B, Rutschow H, Friso G, Rudella A, Emanuelsson O, Sun Q, van Wijk KJ (2008) Sorting signals, N-terminal modifications and abundance of the chloroplast proteome. PLoS ONE 3(4):e1994CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Society for Plant Biochemistry and Biotechnology 2017

Authors and Affiliations

  1. 1.ICAR-National Research Centre on Plant BiotechnologyNew DelhiIndia
  2. 2.School of Life SciencesIndoreIndia

Personalised recommendations