biologia plantarum

International journal on Plant Life established by Bohumil Němec in 1959

Biologia plantarum 62:786-792, 2018 | DOI: 10.1007/s10535-018-0820-5

Molecular cloning and characterization of a PR-5 like protein gene from Brassica campestris ssp. chinensis

C. Liu1,2, H. L. Liu1,2, Y. Wang1,2, D. Hu1,2, D. Xiao1,2, C. W. Zhang1,2, X. L. Hou1,2, Y. Li1,2,*
1 College of Horticulture, Nanjing Agricultural University, Nanjing, P.R. China
2 State Key Laboratory of Crop Genetics and Germplasm Enhancement, Nanjing, P.R. China

Downy mildew caused by Hyaloperonospora parasitica is a serious fungal disease in non-heading Chinese cabbage (Brassica campestris L. ssp. chinensis Makino). Pathogenesis-related 5 (PR-5) genes play an important role in plant resistance to disease invasion. In this study, a gene encoding pathogenesis-related 5-like (PR-5L) protein, named BcPR-5L, was successfully cloned from non-heading Chinese cabbage. The cDNA sequence of BcPR-5L is 747 bp in length. It encoded a protein of molecular mass of 25.78 kDa, an isoelectric point of 4.42, and containing 248 amino acids. Multiple sequence alignment indicated that BcPR-5L protein was highly homologous to other PR-5L proteins identified in 13 different species, with the highest homology to Brassica rapa. We analyzed the subcellular localization of BcPR- 5L protein by using onion epidermal cells and found that it is localized in the membrane. Real time quantitative PCR analyses revealed that the expression of BcPR-5L gene was significantly upregulated after H. parasitica infection, and the expression in the resistant cultivar was higher than that in the susceptible cultivar. In summary, our data suggest that BcPR-5L gene may play an important role in the resistance of non-heading Chinese cabbage to H. parasitica infection.

Keywords: Hyaloperonospora parasitica; non-heading Chinese cabbage; onion; subcellular localization
Subjects: pathogenesis related gene; pathogenesis-related proteins; subcellular localization; amino acid sequences; phylogenetic tree; Chinese cabbage

Received: September 27, 2017; Revised: April 17, 2018; Accepted: April 25, 2018; Published: August 1, 2018  Show citation

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Liu, C., Liu, H.L., Wang, Y., Hu, D., Xiao, D., Zhang, C.W., Hou, X.L., & Li, Y. (2018). Molecular cloning and characterization of a PR-5 like protein gene from Brassica campestris ssp. chinensis. Biologia plantarum, 62(4), 786-792. doi: 10.1007/s10535-018-0820-5
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    References

    1. Ashok Kumar, H.G., Venkatesh, Y.P.: In silico analyses of structural and allergenicity features of sapodilla (Manilkara zapota) acidic thaumatin-like protein in comparison with allergenic plant TLPs. - Mol. Immunology 57: 119-128, 2014. Go to original source...
    2. Batalia, M.A., Monzingo, A.F., Ernst, S., Roberts, W., Robertus, J.D.: The crystal structure of the antifungal protein zeamatin, a member of the thaumatin-like, PR-5 protein family. - Natur. Struct. Biol. 3: 19-23, 1996. Go to original source...
    3. Bormann, C., Baier, D., Horr, I., Raps, C., Berger, J., Jung, G., Schwarz, H.: Characterization of a novel, antifungal, chitinbinding protein from Streptomyces tendae Tu901 that interferes with growth polarity. - J. Bacteriology 181: 7421-7429, 1999. Go to original source...
    4. Brophy, T.F., Laing, M.D.: Screening of fungicides for the control of downy mildew on container-grown cabbage seedlings. - Crop Protect. 11: 160-164, 1992. Go to original source...
    5. Christensen, A.B., Cho, B.H., Naesby, M., Gregersen, P.L., Brandt, J., Madri-Ordenana, K., Collinge, D.B., Thordal-Christensen, H.: The molecular characterization of two barley proteins establishes the novel PR-17 family of pathogenesis-related proteins. - Mol. Plant Pathol. 3: 135-144, 2010. Go to original source...
    6. Datta, K., Velazhahan, R., Oliva, N., Ona, I., Mew, T., Khush, G.S., Muthukrishnan, S., Datta, S.K.: Over-expression of the cloned rice thaumatin-like protein (PR-5) gene in transgenic rice plants enhances environmental friendly resistance to Rhizoctonia solani causing sheath blight disease. - Theor. appl. Genet. 98: 1138-1145, 1999. Go to original source...
    7. Fierens, E., Rombouts, S., Gebruers, K., Goesaert, H., Brijs, K., Beaugrand, J., Volckaert, G., Van Campenhout, S., Proost, P., Courtin, C.M., Delcour, J.A.: TLXI, a novel type of xylanase inhibitor from wheat (Triticum aestivum) belonging to the thaumatin family. - Biochem. J. 403: 583-591, 2007. Go to original source...
    8. Franco, S.F., Baroni, R.M., Carazzolle, M.F., Teixeira, P.J., Reis, O., Pereira, G.A., Mondego, J.M.: Genomic analyses and expression evaluation of thaumatin-like gene family in the cacao fungal pathogen Moniliophthor aperniciosa. - Biochem. biophys. Res. Commun. 466: 629-636, 2015. Go to original source...
    9. Frendo, P., Didierjean, J., Passelegue, E., Burkard, G.: Abiotic stresses induce a thaumatin-like protein in maize; cDNA isolation and sequence analysis. - Plant Sci. 85: 61-69, 1992. Go to original source...
    10. Hu, X., Reddy, A.S.: Cloning and expression of a PR-5-like protein from Arabidopsis: inhibition of fungal growth by bacterially expressed protein. - Plant Mol. Biol. 34: 949-959, 1997. Go to original source...
    11. Jensen, B.D., Hockenhull, J., Munk, L.: Seedling and adult plant resistance to downy mildew (Peronospora parasitica) in cauliflower (Brassica oleracea convar. botrytis var. botrytis). - Plant Pathol. 48: 604-612, 2010. Go to original source...
    12. Kang, Z., Huang, L., Buchenauer, H.: Ultrastructural changes and localization of lignin and callose in compatible and incompatible interactions between wheat and Puccinia striiformis. - J. Plant Dis. Prot. 109: 25-37, 2002.
    13. Liu, K., Zhu, Y., Hou, X., Zhang, S., Cao, S.: [Studies on breeding for disease resistance in non-heading Chinese cabbage IV. Identification and screening for multiple resistance to turnip mosaic virosis, downy mildew and black spot in non-heading Chinese cabbage.] - J. Nanjing Agr. Univ. 20: 31-35, 1997. [In Chinese]
    14. Livak, K.J., Schmittgen, T.D.: Analysis of relative gene expression data using real-time quantitative PCR and the 2 (- Delta Delta C(T)) method. - Methods 25: 402-408, 2001. Go to original source...
    15. Osmond, R.I., Hrmova, M., Fontaine, F., Imberty, A., Fincher, G.B.: Binding interactions between barley thaumatin-like proteins and (1,3)-β-D-glucans. - Eur. J. Biochem. 268: 4190-4199, 2010. Go to original source...
    16. Ovidiu, C., Jamshid, F.: Peronospora-like fungi (Chromista, Peronosporales) parasitic on Brassicaceae and related hosts. - Nova Hedwigia 74: 291-338, 2002. Go to original source...
    17. Parker, J.E., Coleman, M.J., Szabò, V., Frost, L.N., Schmidt, R., Van der Biezen, E.A., Moores, T., Dean, C., Daniels, M.J., Jones, J.D.: The Arabidopsis downy mildew resistance gene RPP5 shares similarity to the toll and interleukin-1 receptors with N and L6. - Plant Cell 9: 879-894, 1997. Go to original source...
    18. Perri, F., Penna, S.D., Rufini, F., Patamia, M., Bonito, M.: Antifungal-protein production in maize (Zea mays) suspension cultures. - Biotechnol. appl. Biochem. 52: 273-281, 2011. Go to original source...
    19. Pierpoint, W.S., Gordonweeks, R., Jackson, P.J.: The occurrence of the thaumatin-like, pathogenesis-related protein, PR-5, in intercellular fluids from Nicotiana species and from an interspecific Nicotiana hybrid. - Physiol. mol. Plant Pathol. 41: 1-10, 1992. Go to original source...
    20. Selitrennikoff, C.P.: Antifungal proteins. - Appl. Environ. Microbiol. 67: 2883-2894, 2001. Go to original source...
    21. Shi, Y.: Mechanisms of caspase activation and inhibition during apoptosis. - Mol. Cell 12: 459-470, 2002. Go to original source...
    22. Sun, C., Wang, L., Hu, D., Riquicho, A.R., Liu, T., Hou, X., Li, Y.: Proteomic analysis of non-heading Chinese cabbage infected with Hyaloperonospora parasitica. - J. Proteomics 98: 15-30, 2014. Go to original source...
    23. Velazhahan, R., Muthukrishnan, S.: Transgenic tobacco plants constitutively overexpressing a rice thaumatin-like protein (PR-5) show enhanced resistance to Alternaria alternata. - Biol. Plant. 47: 347-354, 2003. Go to original source...
    24. Vishunavat, K., Nashaat, N.I., Heran, A., Kolte, S.J.: Sensitivity to the racemic mixture and isomeric forms of metalaxyl in Indian and European homothallic and heterothallic isolates of Peronospora parasitica in Brassica species. - Crop Protect. 17: 543-546, 1998. Go to original source...
    25. Wang, X., Tang, C., Deng, L., Cai, G., Liu, X., Liu, B., Han, Q., Buchenauer, H., Wei, G., Han, D., Huang, L., Kang, Z.: Characterization of a pathogenesis-related thaumatin-like protein gene TaPR5 from wheat induced by stripe rust fungus. - Physiol. Plant. 139: 27-38, 2010. Go to original source...
    26. Xiao, D., Liu, S.T., Wei, Y.P., Zhou, D.Y., Hou, X.L., Li, Y., Hu, C.M.: cDNA-AFLP analysis reveals differential gene expression in incompatible interaction between infected nonheading Chinese cabbage and Hyaloperonospora parasitica. - Hort. Res. 3: 16034, 2016. Go to original source...
    27. Yang, G., Cao, X., Fang, J.G., Song, C.N., Wang, C., Wang, X.C.: Cloning, subcellular localization and spatiotemporal expression of a VvGAI gene from grapevine 'Fujiminori'. - Acta hort. sin. 38: 1883-1892, 2011.
    28. Yerkes, W.D., Shaw, C.G.: Taxonomy of the Peronospora species on Cruciferae and Chenopodiaceae. - Phytopathology 49: 499-507, 1959.

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