Transcriptional Profiles of Mdwrky33 in Apple Root in Response to Infection by Pythium Ultimum, Abiotic Stresses and Chemical Treatments

Yanmin Zhu, Melody Saltzgiver


Plant resistance responses to pathogen infection involve massive transcriptional reprograming and widespread redirection of cellular pathways to adjust the plant from growth to defense. Transcription factors (TFs) function at the major regulating points of gene expression, and specific TFs are known to play crucial roles in plant defense activation. Molecular defense activation in apple root from infection by oomycete necrotrophic pathogen Pythium ultimum, a primary component in a pathogen complex inciting apple replant disease, has not been investigated in detail. Base on previous transcriptome analyses, members of apple WRKY gene family have been identified as the primary candidates in regulating defense response in apple root. Among them, MdWRKY33, an orthologue of AtWRKY33 in apple genome, demonstrated as a highly-expressed WRKY with genotype-specific induction patterns during P. ultimum infection. The sequence features of MdWRKY33 and its tissue-specific expression, as well as its responses to abiotic and pharmacological treatments, added to the evidence for its functional roles in defense activation in apple root. In response to P. ultimum infection, MdWRKY33 was consistently upregulated in all eight tested apple rootstock genotypes at all timepoints. Between genotypes, the stronger induction patterns at the earlier stage of infection in resistant genotypes suggest its essential roles of contributing to apple root resistance, although plant resistance to necrotrophic pathogens is polygenetic quantitative resistance in nature. Transgenic manipulation of this gene is underway to provide more definitive functional identity in contributing to apple root resistance to P. ultimum infection


Apple rootstock; Replant disease; Defense activation; Resistance responses; Transcriptome profiling; Transcription factor; Gene expression; Pythium ultimum; Necrosis patterns


Alves, M. S., S. P. Dadalto, A. B. Goncalves, G. B. de Souza, V. A. Barros and L. G. Fietto. 2014. Transcription Factor Functional Protein-Protein Interactions in Plant Defense Responses. Proteomes, 2: 85-106.

Andreasson, E., T. Jenkins, P. Brodersen, S. Thorgrimsen, N. H. Petersen, S. Zhu, J. L. Qiu, P. Micheelsen, A. Rocher and M. Petersen. 2005. The MAP kinase substrate MKS1 is a regulator of plant defense responses. The EMBO Journal, 24: 2579-89.

Baccelli, I. and B. Mauch-Mani. 2016. Beta-aminobutyric acid priming of plant defense: the role of ABA and other hormones. Plant Molecular Biology, 91: 703-11.

Bakshi, M. and R. Oelmüller. 2014. WRKY transcription factors: Jack of many trades in plants. Plant Signaling & Behavior, 9: e27700.

Benfey, P. N. 2012. Toward a systems analysis of the root Cold Spring Harbor Symposia on Quantitative Biology. pp. 91-96.

Birkenbihl, R. P., C. Diezel and I. E. Somssich. 2012. Arabidopsis WRKY33 is a key transcriptional regulator of hormonal and metabolic responses toward Botrytis cinerea infection. Plant Physiol, 159: 266-85.

Birkenbihl, R. P., S. Liu and I. E. Somssich. 2017. Transcriptional events defining plant immune responses. Current Opinion in Plant Biology, 38: 1-9.

Buscaill, P. and S. Rivas. 2014. Transcriptional control of plant defence responses. Current opinion in plant Biology, 20: 35-46.

Chen, T., W. Li, X. Hu, J. Guo, A. Liu and B. Zhang. 2015. A Cotton MYB Transcription Factor, GbMYB5, is Positively Involved in Plant Adaptive Response to Drought Stress. Plant Cell Physiol, 56: 917-29.

Chezem, W. R. and N. K. Clay. 2016. Regulation of plant secondary metabolism and associated specialized cell development by MYBs and bHLHs. Phytochemistry, 131: 26-43.

Daccord, N., J.-M. Celton, G. Linsmith, C. Becker, N. Choisne, E. Schijlen, H. van de Geest, L. Bianco, D. Micheletti and R. Velasco. 2017. High-quality de novo assembly of the apple genome and methylome dynamics of early fruit development. Nature Genetics, 49: 1099.

Datta, R., D. Kumar, A. Sultana, S. Hazra, D. Bhattacharyya and S. Chattopadhyay. 2015. Glutathione regulates 1-aminocyclopropane-1-carboxylate synthase transcription via WRKY33 and 1-aminocyclopropane-1-carboxylate oxidase by modulating messenger RNA stability to induce ethylene synthesis during stress. Plant Physiology, 169: 2963-81.

De Coninck, B., P. Timmermans, C. Vos, B. P. A. Cammue and K. Kazan. 2015. What lies beneath: Belowground defense strategies in plants. Trends in Plant Science, 20: 91-101.

Eulgem, T., P. J. Rushton, S. Robatzek and I. E. Somssich. 2000. The WRKY superfamily of plant transcription factors. Trends in plant Science, 5: 199-206.

Ishiguro, S. and K. Nakamura. 1994. Characterization of a cDNA encoding a novel DNA-binding protein, SPF1, that recognizes SP8 sequences in the 5′ upstream regions of genes coding for sporamin and β-amylase from sweet potato. Molecular and General Genetics MGG, 244: 563-71.

Ishihama, N. and H. Yoshioka. 2012. Post-translational regulation of WRKY transcription factors in plant immunity. Current Opinion in Plant Biology, 15: 431-37.

Jensen, M. K. and K. Skriver. 2014. NAC transcription factor gene regulatory and protein-protein interaction networks in plant stress responses and senescence. IUBMB Life.

Jin, J., F. Tian, D.-C. Yang, Y.-Q. Meng, L. Kong, J. Luo and G. Gao. 2016. PlantTFDB 4.0: toward a central hub for transcription factors and regulatory interactions in plants. Nucleic Acids Research: gkw982.

Jin, J., H. Zhang, L. Kong, G. Gao and J. Luo. 2014. planttfdb 3.0: a portal for the functional and evolutionary study of plant transcription factors. Nucleic Acids Research, 42: D1182-D87.

Koen, E., P. Trapet, D. Brulé, A. Kulik, A. Klinguer, L. Atauri-Miranda, R. Meunier-Prest, G. Boni, G. Glauser, B. Mauch-Mani, D. Wendehenne and A. Besson-Bard. 2014. β-Aminobutyric Acid (BABA)-Induced Resistance in Arabidopsis thaliana: Link with Iron Homeostasis. Molecular Plant-Microbe Interactions, 27: 1226-40.

Li, J., G. Brader, T. Kariola and E. Tapio Palva. 2006. WRKY70 modulates the selection of signaling pathways in plant defense. Plant Journal, 46: 477-91.

Liu, S., B. Kracher, J. Ziegler, R. P. Birkenbihl and I. E. Somssich. 2015. Negative regulation of ABA signaling by WRKY33 is critical for Arabidopsis immunity towards Botrytis cinerea 2100. elife, 4: e07295.

Livak, K. J. and T. D. Schmittgen. 2001. Analysis of relative gene expression data using real-time quantitative PCR and the 2−ΔΔCT method. Methods, 25: 402-08.

Lorang, J. 2018. Necrotrophic Exploitation and Subversion of Plant Defense: A Lifestyle or Just a Phase, and Implications in Breeding Resistance. Phytopathology, 109: 332-46.

Mangelsen, E., J. Kilian, K. W. Berendzen, Ü. H. Kolukisaoglu, K. Harter, C. Jansson and D. Wanke. 2008. Phylogenetic and comparative gene expression analysis of barley (Hordeum vulgare) WRKY transcription factor family reveals putatively retained functions between monocots and dicots. BMC Genomics, 9: 194.

Mao, G., X. Meng, Y. Liu, Z. Zheng, Z. Chen and S. Zhang. 2011. Phosphorylation of a WRKY Transcription Factor by Two Pathogen-Responsive MAPKs Drives Phytoalexin Biosynthesis in Arabidopsis. The Plant Cell, 23: 1639-53.

Martinez-Medina, A., V. Flors, M. Heil, B. Mauch-Mani, C. M. J. Pieterse, M. J. Pozo, J. Ton, N. M. van Dam and U. Conrath. 2016. Recognizing Plant Defense Priming. Trends in Plant Science, 21: 818-22.

Mazzola, M. 1998. Elucidation of the microbial complex having a causal role in the development of apple replant disease in washington. Phytopathology, 88: 930-8.

Meng, D., Y. Li, Y. Bai, M. Li and L. Cheng. 2016. Genome-wide identification and characterization of WRKY transcriptional factor family in apple and analysis of their responses to waterlogging and drought stress. Plant Physiology and Biochemistry, 103: 71-83.

Menke, F. L., H.-G. Kang, Z. Chen, J. M. Park, D. Kumar and D. F. Klessig. 2005. Tobacco transcription factor WRKY1 is phosphorylated by the MAP kinase SIPK and mediates HR-like cell death in tobacco. Molecular Plant-Microbe Interactions, 18: 1027-34.

Patra, B., C. Schluttenhofer, Y. Wu, S. Pattanaik and L. Yuan. 2013. Transcriptional regulation of secondary metabolite biosynthesis in plants. Biochimica et Biophysica Acta (BBA) - Gene Regulatory Mechanisms, 1829: 1236-47.

Ren, X., Z. Chen, Y. Liu, H. Zhang, M. Zhang, Q. Liu, X. Hong, J. K. Zhu and Z. Gong. 2010. ABO3, a WRKY transcription factor, mediates plant responses to abscisic acid and drought tolerance in Arabidopsis. Plant J, 63: 417-29.

Riechmann, J. L., J. Heard, G. Martin, L. Reuber, C.-Z. Jiang , J. Keddie, L. Adam, O. Pineda, O. J. Ratcliffe, R. R. Samaha, R. Creelman, M. Pilgrim, P. Broun, J. Z. Zhang, D. Ghandehari, B. K. Sherman and G. -L. Yu. 2000. Arabidopsis Transcription Factors: Genome-Wide Comparative Analysis Among Eukaryotes. Science, 290: 2105-10.

Rushton, P. J., M. T. Bokowiec, S. Han, H. Zhang, J. F. Brannock, X. Chen, T. W. Laudeman and M. P. Timko. 2008. Tobacco Transcription Factors: Novel Insights into Transcriptional Regulation in the Solanaceae. Plant Physiology, 147: 280-95.

Samad, A. F. A., M. Sajad, N. Nazaruddin, I. A. Fauzi, A. M. A. Murad, Z. Zainal and I. Ismail. 2017. MicroRNA and Transcription Factor: Key Players in Plant Regulatory Network. Front Plant Sci, 8: 565.

Shin, S., J. Lee, D. Rudell, K. Evans and Y. Zhu. 2016. Transcriptional regulation of auxin metabolism and ethylene biosynthesis activation during apple (Malus domestica) fruit maturation. Journal of Plant Growth Regulation, 35: 655-66.

Shin, S., P. Zheng, G. Fazio, M. Mazzola, D. Main and Y. Zhu. 2016. Transcriptome changes specifically associated with apple (Malus domestica) root defense response during Pythium ultimum infection. Physiological and Molecular Plant Pathology, 94: 16-26.

Singh, K. B., R. C. Foley and L. Oñate-Sánchez. 2002. Transcription factors in plant defense and stress responses. Current Opinion in Plant Biology, 5: 430-36.

Song, Y., C. Ai, S. Jing and D. Yu. 2010. Research progress on functional analysis of rice WRKY genes. Rice science, 17: 60-72.

Van Verk, M. C., C. Gatz and H. J. Linthorst. 2009. Transcriptional regulation of plant defense responses. Advances in Botanical Research, 51: 397-438.

Velasco, R., A. Zharkikh, J. Affourtit, A. Dhingra, A. Cestaro, A. Kalyanaraman, P. Fontana, S. K. Bhatnagar, M. Troggio and D. Pruss. 2010. The genome of the domesticated apple (Malus domestica Borkh.). Nature Genetics, 42: 833.

Wang, X., N. Jiang, J. Liu, W. Liu and G. L. Wang. 2014. The role of effectors and host immunity in plant-necrotrophic fungal interactions. Virulence, 5: 722-32.

Wasternack, C. and M. Strnad. 2019. Jasmonates are signals in the biosynthesis of secondary metabolites — Pathways, transcription factors and applied aspects — A brief review. New Biotechnology, 48: 1-11.

Wu, K.-L., Z.-J. Guo, H.-H. Wang and J. Li. 2005. The WRKY family of transcription factors in rice and Arabidopsis and their origins. DNA Research, 12: 9-26.

Zhang, H., J. Jin, L. Tang, Y. Zhao, X. Gu, G. Gao and J. Luo. 2011. PlantTFDB 2.0: update and improvement of the comprehensive plant transcription factor database. Nucleic Acids Res, 39: D1114-7.

Zhang, Y., H. Yu, X. Yang, Q. Li, J. Ling, H. Wang, X. Gu, S. Huang and W. Jiang. 2016. CsWRKY46, a WRKY transcription factor from cucumber, confers cold resistance in transgenic-plant by regulating a set of cold-stress responsive genes in an ABA-dependent manner. Plant Physiol Biochem, 108: 478-87.

Zhou, M. and J. Memelink. 2016. Jasmonate-responsive transcription factors regulating plant secondary metabolism. Biotechnology Advances, 34: 441-49.

Zhu, Y., M. Saltzgiver and J. Zhao. 2018. A Phenotyping Protocol for Detailed Evaluation of Apple Root Resistance Responses Utilizing Tissue Culture Micropropagated Apple Plants. American Journal of Plant Sciences, 9: 2183.

Zhu, Y., J. Shao, Z. Zhou and R. E. Davis. 2017. Comparative Transcriptome Analysis Reveals a Preformed Defense System in Apple Root of a Resistant Genotype of G.935 in the Absence of Pathogen. International Journal of Plant Genomics, 2017: 8950746.

Zhu, Y., J. Shao, Z. Zhou and R. E. Davis. 2019. Genotype-specific suppression of multiple defense pathways in apple root during infection by Pythium ultimum. Horticulture Research, 6: 10-16.

Zhu, Y., S. Shin and M. Mazzola. 2016. Genotype responses of two apple rootstocks to infection by Pythium ultimum causing apple replant disease. Canadian Journal of Plant Pathology, 38: 483-91.

Zhu, Y., J. Zhao and Z. Zhou. 2018. Identifying an Elite Panel of Apple Rootstock Germplasm with Contrasting Root Resistance to Pythium ultimum. Journal of Plant Pathology & Microbiology, 9:34-39.

Full Text: PDF

DOI: 10.33687/phytopath.008.03.2996


  • There are currently no refbacks.

Copyright (c) 2019 Yanmin Zhu

Creative Commons License
This work is licensed under a Creative Commons Attribution 4.0 International License.