biologia plantarum

International journal on Plant Life established by Bohumil Nìmec in 1959

Biologia plantarum 61:763-773, 2017 | DOI: 10.1007/s10535-017-0726-7

Isolation and molecular characterization of pathogenesis related PR2 gene and its promoter from Brassica juncea

S. Ali1,2, N. Chandrashekar3, S. Rawat1, N. M. C. Nayanakantha1, Z. A. Mir1, A. Manoharan4, M. Sultana2, A. Grover1,*
1 National Research Centre on Plant Biotechnology, Pusa Campus, New Delhi, India
2 Department of Advanced Zoology and Biotechnology, Presidency College, Chennai, India
3 Division of Microbiology, CCUBGA, Indian Agricultural Research Institute, New Delhi, India
4 Department of Plant Biology and Biotechnology, Presidency College, Chennai, India

Pathogenesis-related (PR) proteins play key roles in plant disease resistance. Here, we isolated and characterized pathogenesis-related PR2 gene encoding β-1,3-glucanase from Brassica juncea and named it BjPR2 (GenBank accession number DQ359125). The amino acid sequence of BjPR2 showed ~99 % similarity with β-1,3-glucanase of Brassica rapa, B. napus, and B. oleracea. BjPR2 transcription was rapidly increased after Alternaria brassicae infection, salicylic acid application, and wounding, but the induction was delayed in response to jasmonic acid. To investigate the transcriptional regulation of BjPR2 gene, its promoter was isolated. In silico analysis of BjPR2 promoter showed cis-regulatory elements upstream of TATA and CAAT boxes responsive to defense, hormones, wounding, and plant developmental stage. Homozygous Arabidopsis thaliana lines were developed with plasmid construct having β-glucuronidase (GUS) reporter gene driven by BjPR2 promoter. The analysis of GUS protein in Arabidopsis lines showed that BjPR2 promoter drived distinct pattern of pathogen inducible expression after fungal infection (Alternaria brassicae, Erysiphe orontii), phytohormones, and wounding. It also showed age dependent and organ specific expressions. BjPR2 promoter drove strong GUS activity in Arabidopsis seedlings and showed organ specific expression at the later growth stages (lateral organ junctions, leaf serrate, base of siliques, and receptacle). Due to stress-inducible and tissue specific nature, the BjPR2 promoter can serve as a potential candidate in genetic engineering.

Keywords: Alternaria brassicae; Arabidopsis thaliana; β-glucuronidase; Indian mustard; jasmonic acid; phylogenetic tree; salicylic acid; transgenic plant
Subjects: pathogenesis related gene; β-glucuronidase; amino acid sequences; phylogenetic tree; jasmonic acid; salicylic acid; wounding; transgenic plants; Indian mustard; fungal infection

Received: August 31, 2016; Revised: January 18, 2017; Accepted: January 27, 2017; Published: December 1, 2017  Show citation

ACS AIP APA ASA Harvard Chicago IEEE ISO690 MLA NLM Turabian Vancouver
Ali, S., Chandrashekar, N., Rawat, S., Nayanakantha, N.M.C., Mir, Z.A., Manoharan, A., Sultana, M., & Grover, A. (2017). Isolation and molecular characterization of pathogenesis related PR2 gene and its promoter from Brassica juncea. Biologia plantarum, 61(4), 763-773. doi: 10.1007/s10535-017-0726-7
Share...
Download citation
  • BibTeX (.bib)
  • Bookends (.ris)
  • EasyBib (.ris)
  • EndNote (.enw)
  • EndNote 8 (.xml)
  • ISI WoS (.isi)
  • Medlars (.medlars)
  • Mendeley (.ris)
  • MODS (.xml)
  • Papers (.ris)
  • RefWorks (.txt)
  • RefManager (.ris)
  • RIS (.ris)
  • MS Word (.xml)
  • Zotero (.ris)
    • Open full article

    Supplementary files

    Download filebpl-201704-0019_S1.pdf

    File size: 130.58 kB

    References

    1. Aggarwal, R., Purwar, S., Kharbikar, L., Sangeeta Gupta, L.: Induction of a wheat ß-1,3-glucanase gene during the defense response to Bipolaris sorokiniana. - Acta phytopathol. entomol. hung. 46: 37-49, 2011. Go to original source...
    2. Akiyama, T., Jin, S., Yoshida, M., Hoshino, T., Opaßsiri, R., Ketudat Cairns, J.R.: Expression of an endo-(1,3;1,4) β-glucanase in response to wounding, methyl jasmonate, abscisic acid and ethephon in rice seedlings. - J. Plant Physiol. 166: 1814-1825, 2009. Go to original source...
    3. Al-Taweel, K., Fernando, W.G.D., Brule-Babel, A.L.: Construction and characterization of a cDNA library from wheat infected with Fusarium graminearum Fg 2. - Int. J. mol. Sci. 12: 613-626, 2011. Go to original source...
    4. Balbi, V., Devoto, A.: Jasmonate signaling network in Arabidopsis thaliana: crucial regulatory nodes and new physiological scenarios. - New Phytol. 177: 301-318, 2008. Go to original source...
    5. Cheong, Y.H., Kim, C.Y., Chun, H.J., Moon, B.C., Park, H.C., Kim, J.K., Lee, S.H., Han, C.D., Lee, S.Y., Cho, M.J.: Molecular cloning of a soybean claßs III β-1,3-glucanase gene that is regulated both developmentally and in response to pathogen infection. - Plant Sci. 154: 71-81, 2000. Go to original source...
    6. Clough, S.J., Bent, A.: Floral dip: a simplified method for Agrobacterium-mediated transformation of Arabidopsis thaliana. - Plant J. 16: 735-743, 1998. Go to original source...
    7. Durrant, W.E., Rowland, O., Piedras, P., Hammond-Kosack, K.E., Jones, J.D.G.: cDNA-AFLP reveals a striking overlap in race-specific resistance and wound response gene expression profiles. - Plant Cell 12: 963-977, 2000. Go to original source...
    8. Eulgem, T., Rushton, P.J., Robatzek, S., Somssich, I.E.: The WRKY super family of plant transcription factors. - Trends Plant Sci. 5: 199-206, 2000. Go to original source...
    9. Filichkin, S.A., Leonard, J.M., Monteros, A., Liu, P.P., Nonogaki, H.: A novel endo-beta-mannanase gene in tomato LeMAN5 is associated with anther and pollen development. - Plant Physiol. 134: 1080-1087, 2004. Go to original source...
    10. Fung, R.W.M., Gonzalo, M., Fekete, C., Kovacs, L.G., He, Y., Marsh, E., McIntyre, L.M., Schachtman, D.P., Qiu, W.P.: Powdery mildew induces defense oriented reprogramming of the transcriptome in a susceptible but not in a resistant grapevine. - Plant Physiol. 146: 236-249, 2008. Go to original source...
    11. Grover, A., Gowthaman, R.: Strategies for development of fungus resistant transgenic plants. - Curr. Sci. 84: 330-340, 2003.
    12. Gupta, P., Ravi, I., Sharma, V.: Induction of ß-1,3-glucanase and chitinase activity in the defense response of Eruca sativa plants against the fungal pathogen Alternaria brassicicola. - J. Plant. Interact. 8: 155-161, 2013. Go to original source...
    13. Higo, K., Ugawa, Y., Iwamoto, M., Korenaga, T.: Plant cis-acting regulatory DNA elements (PLACE) database. - Nucl. Acids Res. 27:297-300, 1999. Go to original source...
    14. Jayaraj, J., Muthukrishnan, S., Liang, G.H., Velazhahan, R.: JA and SA induce accumulation of ß-1,3-glucanase and thaumatin-like proteins in wheat and enhance resistance against Stagonospora nodorum. - Biol. Plant. 48: 425-430, 2004. Go to original source...
    15. Jefferson, R.A., Kavanagh, T.A., Bevan, M.W.: GUS fusions: beta-glucuronidase as a sensitive and versatile gene fusion marker in higher plants. - EMBO J. 6: 3901-3907, 1987. Go to original source...
    16. Jorda, L., Vera, P.: Local and systemic induction of two defense-related subtilisin-like protease promoters in transgenic Arabidopsis plants. Luciferin induction of PR gene expression. - Plant Physiol. 124: 1049-1057, 2000. Go to original source...
    17. Ko, T.S., Lee, S., Schaefer, S.C., Korban, S.S.: Characterization of a tißsue-specific and developmentally regulated β-1,3-glucanase gene family in Prunus persica. - Plant Physiol. Biochem. 41: 955-963, 2003. Go to original source...
    18. Kumar, S., Stecher, G., Tamura, K.: MEGA7: molecular evolutionary genetics analysis version 7.0 for bigger datasets. - Mol. Biol. Evol. doi: 10.1093/molbev/msw054, 2016. Go to original source...
    19. Lescot, M., Dehais, P., Thijs, G., Marchal, K., Moreau, Y., Van De Peer, Y., Rouze, P., Rombauts, S.: PlantCARE, a database of plant cis-acting regulatory elements and a portal to tools for in silico analysis of promoter sequences. - Nucl. Acids Res. 30: 325-327, 2002. Go to original source...
    20. Li, Y.F., Zhu, R., Xu, P.: Activation of the gene promoter of barley ß-1,3-glucanase isoenzymes GIII is SA (SA)- dependent in transgenic rice plants. - J. Plant Res. 118: 215-221, 2005. Go to original source...
    21. Liu, B., Xue, X.D., Cui, S.P., Zhang, X.Y., Han, Q.M., Zhu, L., Liang, X.F., Wang, X.J., Huang, L.L., Chen, X.M., Kang, Z.S.: Cloning and characterization of a wheat ß-1,3-glucanase gene induced by the stripe rust pathogen Puccinia striiformis f. sp. tritici. - Mol. Biol. Rep. 37: 1045-1052, 2010. Go to original source...
    22. Livak, K.J., Schmittgen, T.D.: Analysis of relative gene expression data using real-time quantitative PCR and the 2-ΔΔCt method. - Methods 25: 402-408, 2001. Go to original source...
    23. Maleck, K., Dietrich, R.A.: Defense on multiple fronts: how do plants cope with diverse enemies? - Trends Plant. Sci. 4: 215-219, 1999. Go to original source...
    24. Mazarei, M., Teplova, I., Hajimorad, M.R., Stewart, C.N.: Pathogen phytosensing: plants to report plant pathogens. - Sensors 8: 2628-2641, 2008. Go to original source...
    25. Mondal, K.K., Bhattacharya, R.C., Koundal, K.R., Chatterjee, S.C.: Transgenic Indian mustard (Brassica juncea) expressing tomato glucanase leads to arrested growth of Alternaria brassicae. - Plant Cell Rep. 26: 247-252, 2007. Go to original source...
    26. Murashige, T., Skoog, T.: A revised medium for rapid growth and bioassays with tobacco tissue cultures. - Physiol. Plant. 15: 473-497, 1962. Go to original source...
    27. Ohme-Takagi, M., Shinshi, H.: Ethylene-inducible DNA binding proteins that interact with an ethylene-responsive element. - Plant Cell 7: 173-182, 1995. Go to original source...
    28. Otani, H., Kohnobe, A., Kodama, M., Kohmoto, K.: Production of a host-specific toxin by germinating spores of Alternaria brassicicola. - Physiol. mol. Plant Pathol. 52: 285-295, 1998. Go to original source...
    29. Reuber, T.L., Plotnikova, J.M., Dewdney, J., Rogers, E.E., Wood, W., Ausubel, F.M.: Correlation of defense gene induction defects with powdery mildew susceptibility in Arabidopsis enhanced disease susceptibility mutants. - Plant J. 16: 473-485, 1998. Go to original source...
    30. Rogers, H.J., Bate, N., Combe, J., Sullivan, J., Sweetman, J., Swan, C., Lonsdale, D.M., Twell, D.: Functional analysis of cis-regulatory elements within the promoter of the tobacco late pollen gene g10. - Plant. mol. Biol. 45: 577-585, 2001. Go to original source...
    31. Rojo, E., Leon, J., Sanchez-Serrano, J.J.: Cross-talk between wound signaling pathways determines local versus systemic gene expression in Arabidopsis thaliana. - Plant J. 20: 135-142, 1999. Go to original source...
    32. Rushton, P.J., Somssich, I.E.: Transcriptional control of plant genes responsive to pathogens. - Curr. Opin. Plant. Biol. 1: 311-315, 1998. Go to original source...
    33. Sa, Q., Wang, Y., Li, W., Zhang, L., Sun, Y.: The promoter of an antifungal protein gene from Gastrodia elata confers tissue-specific and fungus-inducible expression patterns and responds to both salicylic acid and jasmonic acid. - Plant. Cell. Rep. 22: 79-84, 2003. Go to original source...
    34. Seo, P.J., Lee, A.K., Xiang, F., Park, C.M.: Molecular and functional profiling of Arabidopsis pathogenesis-related genes: insights into their roles in salt response of seed germination. - Plant Cell Physiol. 49: 334-344, 2008. Go to original source...
    35. Shi, Y.L., Zhang, Y.H., Shih, DS.: Cloning and expreßsion analysis of two β-1, 3 glucanase gene from strawberry. - J. Plant Physiol. 163: 956-967, 2006. Go to original source...
    36. Siebert, P.D., Chenchik, A., Kellogg, D.E., Lukyanov, K.A., Lukyanov, S.A.: An improved PCR method for walking in uncloned genomic DNA. - Nucl. Acids Res. 23:1087-1088, 1995. Go to original source...
    37. Thakur, R., Kolte, S.J.: Radish root extract agar, a suitable medium for the growth and sporulation of Alternaria brassicae. - Cruciferae News Lett. 10: 117-118, 1985.
    38. Thomma, B.P.H.J., Eggermont, K., Penninckx, I.A.M.A., Mauch Mani, B., Vogelsang, R., Cammue, B.P.A., Broekaert, W.F.: Separate jasmonate-dependent and salicylate dependent defense-response pathways in Arabidopsis are essential for resistance to distinct microbial pathogens. - Proc. nat. Acad. Sci. USA 95: 15107-15111, 1998. Go to original source...
    39. Thompson, J.D., Gibson, T.J., Plewniak, F., Jeanmougin, F., Higgins, D.G.: The ClustalX windows interface: flexible strategies for multiple sequence alignment aided by quality analysis tools. - Nucl. Acids Res. 25: 4876-4882, 1997. Go to original source...
    40. Vinu, V., Singh, N., Vasudev, S., Yadava, D.K., Kumar, S., Naresh, S., Bhat, S.R., Prabhu, K.: Assessment of genetic diversity in Brassica juncea (Brassicaceae) genotypes using phenotypic differences and SSR markers. - Rev. Biol. Trop. 61: 1919-1934, 2013.
    41. Vishwanath, K., Kolte, S.J.: Variability in Alternaria brassicae: response to host genotypes, toxin production and fungicides. - Indian Phytopathol. 50: 373-381, 1997.
    42. Wan, L., Zha, W., Cheng, X., Liu, C., Lv, L., Liu, C., Wang, Z., Du, B., Chen, R., Zhu, L., He, GA.: Rice ß-1,3-glucanase gene Osg1 is required for callose degradation in pollen development. - Planta 233: 309-323, 2011. Go to original source...
    43. Wang, W., Barnaby, J.Y., Tada, Y., Li, H., Tör, M., Caldelari, D., Lee, D.U., Fu, X.D., Dong, X.N.: Timing of plant immune responses by a central circadian regulator. - Nature 470: 110-114, 2011. Go to original source...
    44. Wu, C.T., Bradford, K.J.: Claßs I chitinase and β-1,3-glucanase are differentially regulated by wounding, methyl jasmonate, ethylene, and gibberellin in tomato seeds and leaves. - Plant Physiol. 133: 263-273, 2003. Go to original source...
    45. Yadava, J.S., Singh, N.B.: Strategies to enhance yield potential of rapeseed-mustard in India. - In: Wratten, N., Salisbury, P. (ed.): Proc. 10th International Rapeseed Congress. Pp. 26-29. Canberra 1999.
    46. Yang, Y.X., Wang, M.M., Yin, Y.L., Onac, E., Zhou, G.F., Peng, S.: RNA-seq analysis reveals the role of red light in resistance against Pseudomonas syringae pv. tomato DC3000 in tomato plants. - BMC Genomics 16: 120, 2015. Go to original source...
    47. Yin, L.L., Hou, X.J.: The recent advances of salicylic acid as signal molecules of resistance in plant. - Chin. Agr. Sci Bull. 123: 338-342. 2007.
    48. Zamora, M.G.M., Bournonville, C.G., Castagnaro, A.P., Ricci, J.C.D.: Identification and characterization of a novel claßs I endo-β-1,3-glucanase regulated by SA, ethylene and fungal pathogens in strawberry. - Funct. Plant Biol. 39: 412-420, 2012. Go to original source...
    49. Zemanek, A.B., Ko, T.S., Thimmapuram, J., Hammerschlag, F.A., Korban, S.S.: Changes in ß-1,3-glucanase mRNA levels in peach in response to treatment with pathogen culture filtrates, wounding and other elicitors. - J. Plant Physiol. 159: 877-889, 2002. Go to original source...
    50. Zheng, X., Deng, W., Luo, K., Duan, H., Chen, Y., McAvoy, R., Song, S., Pei, Y., Li, Y.: The cauliflower mosaic virus (CaMV) 35S promoter sequence alters the level and patterns of activity of adjacent tissue and organ-specific gene promoters. - Plant. Cell Rep. 26: 1195-1203, 2007. Go to original source...

    Return to the content


    -