Tree Genetics & Genomes

, 14:75 | Cite as

RNA sequencing analysis provides new insights into dynamic molecular responses to Valsa mali pathogenicity in apple ‘Changfu No. 2’

  • Cunwu Zuo
  • Juan Mao
  • Zhongjian Chen
  • Mingyu Chu
  • Hu Duo
  • Baihong ChenEmail author
Original Article
Part of the following topical collections:
  1. Disease Resistance


Valsa canker caused by the necrotrophic pathogen Valsa mali (Vm) severely affects apple production in Eastern Asia. The molecular basis underlying the apple response to Vm infection is poorly understood. Hence, we performed RNA sequencing (RNA-seq) to investigate the dynamic gene expression profiles of a major apple cultivar, ‘Changfu No.2’, during Vm infection. Compared with the control (C), 104, 313, and 1059 differentially expressed genes (DEGs) were detected from the phloem tissue within the range of 0.9–1.3 cm (T1), 0.5–0.9 cm (T2), and 0.1–0.5 cm (T3) beyond the lesion periphery, respectively. Gene ontology (GO) enrichment analysis revealed that the DEGs associated with plant growth and development were down-regulated, whereas those related to defense responses were up-regulated. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis showed that hormonal and Ca2+ signaling and phenylpropanoid biosynthesis were involved in the defense responses. In conclusion, multiple defense responses associated with ABA, JA, ET, Ca2+, and cell wall signals contributed to the defense against Vm infection in ‘Changfu No.2’. In contrast, the DEGs with inhibited expression were involved in plant growth and development; auxin signaling and several resistance genes might weaken the resistance of ‘Changfu No.2’ to pathogens. Our results offer a new insight into plant responses against necrotrophs and could benefit programs aimed at breeding for Vm resistance.


Valsa canker Apple Hormone Ca2+-signal Defense responses 



Azelaic acid


Calcium ion


Cell wall degrading enzymes


Differentially expressed genes




Effector-triggered immunity






Jasmonic acid


Methyl salicylic acid


Pathogen-associated molecular pattern


Pattern-recognition receptor


PAMP-triggered immunity


Real-time quantitative PCR


RNA sequencing


Salicylic acid


Systemic acquired resistance


Transcription factor


Valsa mali



We would like to thank Ph.D Lijun Bai (GeneBang Inc., Chengdu, China, for technical assistance with RNA sequencing and bioinformatic analysis.

Author’s contributions

ZC and CB conceived, designed, and coordinated the study. ZC, MJ, CM, and DH performed the experiments and collected, analyzed, and deposited the data. CZ proofread the final draft and revised the manuscript. All authors have read and approved the manuscript.


This work was supported by the Talent introduction Project of Gansu Agricultural University (GSAU-RCZX201712) and the Natural Science Foundation of China No. 31501728.

Compliance with ethical standards

Conflict of interest

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Ethical approval

This article does not contain any studies with human participants performed by any of the authors.

Data Archiving Statement

The raw data has been deposited in the National Center for Biotechnology information (NCBI) Short Read Archive (SPA) under accession number SRP160545.

Supplementary material

11295_2018_1288_MOESM1_ESM.docx (14 kb)
ESM 1 (DOCX 13 kb)


  1. Abe K, Kotoda N, Kato H, Soejima J (2007) Resistance sources to Valsa canker (Valsa ceratosperma) in a germplasm collection of diverse Malus species. Plant Breed 126:449–453CrossRefGoogle Scholar
  2. Bacete L, Mélida H, Miedes E, Molina A (2018) Plant cell wall-mediated immunity: cell wall changes trigger disease resistance responses. Plant J 93:614–636CrossRefPubMedGoogle Scholar
  3. Bellincampi D, Cervone F, Lionetti V (2014) Plant cell wall dynamics and wall-related susceptibility in plant-pathogen interactions. Front Plant Sci 5:228CrossRefPubMedPubMedCentralGoogle Scholar
  4. Berens ML, Berry HM, Mine A, Argueso CT, Tsuda K (2016) Evolution of hormone signaling networks in plant defense. Annu Rev Phytopathol 55:401–425CrossRefGoogle Scholar
  5. Berger S, Sinha AK, Roitsch T (2007) Plant physiology meets phytopathology: plant primary metabolism and plant-pathogen interactions. J Exp Bot 58:4019–4026CrossRefPubMedGoogle Scholar
  6. Berrocal-Lobo M, Molina A (2008) Arabidopsis defense response against Fusarium oxysporum. Trends Plant Sci 13:145–150CrossRefPubMedGoogle Scholar
  7. Cantu D, Vicente AR, Labavitch JM, Bennett AB, Powell AL (2008) Strangers in the matrix: plant cell walls and pathogen susceptibility. Trends Plant Sci 13:610–617CrossRefPubMedGoogle Scholar
  8. Cao K, Guo L, Li B, Sun G, Chen H (2009) Investigations on the occurrence and control of apple canker in China. Plant Prot 35:114–116 (In Chinese)Google Scholar
  9. Chanda B, Xia Y, Mandal MK, Yu K, Sekine KT, Gao QM, Selote D, Hu YL, Stromberg A, Navarre D, Kachroo A, Kachroo P (2011) Glycerol-3-phosphate is a critical mobile inducer of systemic immunity in plants. Nat Genet 43:421–427CrossRefPubMedGoogle Scholar
  10. Chaturvedi R, Venables B, Petros RA, Nalam V, Li M, Wang X, Takemoto LJ, Shah J (2012) An abietane diterpenoid is a potent activator of systemic acquired resistance. Plant J 71:161–172CrossRefPubMedGoogle Scholar
  11. Daccord N, Celton JM, Linsmith G, Becker C, Choisne N, Schijlen E, Geest H, Bianco L, Micheletti D, Velasco R, Pierro E, Gouzy J, Rees D, Guérif P, Muranty H, Durel CE, Laurens F, Lespinasse Y, Gaillard S, Aubourg S, Quesneville H, Weigel D, Weg E, Troggio M, Bucher E (2017) High-quality de novo assembly of the apple genome and methylome dynamics of early fruit development. Nat Genet 49:1099–1106CrossRefPubMedGoogle Scholar
  12. Dörmann P, Kim H, Ott T, Schulze-Lefert P, Trujillo M, Wewer V, Hückelhoven R (2014) Cell-autonomous defense, re-organization and trafficking of membranes in plant-microbe interactions. New Phytol 204:815–822CrossRefPubMedGoogle Scholar
  13. Fu Z, Dong X (2013) Systemic acquired resistance: turning local infection into global defense. Annu Rev Plant Biol 64:839–863CrossRefPubMedGoogle Scholar
  14. Gao Q, Zhu S, Kachroo P, Kachroo A (2015) Signal regulators of systemic acquired resistance. Front Plant Sci 6:228PubMedPubMedCentralGoogle Scholar
  15. Genger RK, Jurkowski GI, McDowell JM, Lu H, Jung HW, Greenberg JT, Bent AF (2008) Signaling pathways that regulate the enhanced disease resistance of Arabidopsis “defense, no death” mutants. Mol Plant Microbe Interact 21:1285–1296CrossRefPubMedPubMedCentralGoogle Scholar
  16. Glazebrook J (2005) Contrasting mechanisms of defense against biotrophic and necrotrophic pathogens. Annu Rev Phytopathol 43:205–227CrossRefPubMedGoogle Scholar
  17. Harwood J (2012) Lipids in plants and microbes. Springer Science & Business Media, BerlinGoogle Scholar
  18. Jones JD, Dangl JL (2006) The plant immune system. Nature 444:323–329CrossRefPubMedGoogle Scholar
  19. Kanehisa M, Araki M, Goto S, Hattori M, Hirakawa M, Itoh M, Katayama T, Kawashima S, Okuda S, Tokimatsu T, Yamanishi Y (2008) KEGG for linking genomes to life and the environment. Nucleic Acids Res 36:D480–D484CrossRefPubMedGoogle Scholar
  20. Kazan K, Manners JM (2009) Linking development to defense: auxin in plant-pathogen interactions. Trends Plant Sci 14:373–382CrossRefPubMedGoogle Scholar
  21. Kepley JB, Jacobi WR (2000) Pathogenicity of Cytospora fungi on six hardwood species. J Arboric 26:326–332Google Scholar
  22. Kidd BN, Kadoo NY, Dombrecht B, Tekeoglu M, Gardiner DM, Thatcher LF, Aitken E, Schenk P, Manners J, Kazan K (2011) Auxin signaling and transport promote susceptibility to the root-infecting fungal pathogen Fusarium oxysporum in Arabidopsis. Mol Plant Microbe Interact 24:733–748CrossRefPubMedGoogle Scholar
  23. Li B, Dewey CN (2011) RSEM: accurate transcript quantification from RNA-Seq data with or without a reference genome. BMC Bioinformatics 12:323CrossRefPubMedPubMedCentralGoogle Scholar
  24. Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative and the 2-ΔΔCt method. Methods 25:402–408CrossRefGoogle Scholar
  25. Llorente F, Alonso-Blanco C, Sánchez-Rodriguez C, Jorda L, Molina A (2005) ERECTA receptor-like kinase and heterotrimeric G protein from Arabidopsis are required for resistance to the necrotrophic fungus Plectosphaerella cucumerina. Plant J 43:165–180CrossRefPubMedGoogle Scholar
  26. Llorente F, Muskett P, Sánchez-Vallet A, López G, Ramos B, Sánchez-Rodríguez C, Jordá L, Parker J, Molina A (2008) Repression of the auxin response pathway increases Arabidopsis susceptibility to necrotrophic fungi. Mol Plant 1:496–509CrossRefPubMedGoogle Scholar
  27. López-Berges MS, Rispail N, Prados-Rosales RC, Di Pietro A (2010) A nitrogen response pathway regulates virulence functions in Fusarium oxysporum via the protein kinase TOR and the bZIP protein MeaB. Plant Cell 22:2459–2475CrossRefPubMedPubMedCentralGoogle Scholar
  28. Magnin-Robert M, Le BD, Markham J, Dorey S, Clément C, Baillieul F, Dhondt-Cordelier S (2015) Modifications of sphingolipid content affect tolerance to hemibiotrophic and necrotrophic pathogens by modulating plant defense responses in arabidopsis. Plant Physiol 169:2255–2274PubMedPubMedCentralGoogle Scholar
  29. Mengiste T (2012) Plant immunity to necrotrophs. Annu Rev Phytopathol 50:267–294CrossRefPubMedGoogle Scholar
  30. Mengiste T, Chen X, Salmeron J, Dietrich R (2003) The BOTRYTIS SUSCEPTIBLE1 gene encodes an R2R3MYB transcription factor protein that is required for biotic and abiotic stress responses in Arabidopsis. Plant Cell 15:2551–2565CrossRefPubMedPubMedCentralGoogle Scholar
  31. Moeder W, Urquhart W, Ung H, Yoshioka K (2011) The role of cyclic nucleotide-gated ion channels in plant immunity. Mol Plant 4:442–452CrossRefPubMedGoogle Scholar
  32. Naseem M, Srivastava M, Tehseen M, Ahmed N (2015) Auxin crosstalk to plant immune networks: a plant-pathogen interaction perspective. Curr Protein Pept Sci 16:389–394CrossRefPubMedGoogle Scholar
  33. Oliver RP, Ipcho SV (2004) Arabidopsis pathology breathes new life into the necrotrophs-vs.-biotrophs classification of fungal pathogens. Mol Plant Pathol 5:347–352CrossRefPubMedGoogle Scholar
  34. Pré M, Atallah M, Champion A, De Vos M, Pieterse CM, Memelink J (2008) The AP2/ERF domain transcription factor ORA59 integrates jasmonic acid and ethylene signals in plant defense. Plant Physiol 147:1347–1357CrossRefPubMedPubMedCentralGoogle Scholar
  35. Ranty B, Aldon D, Cotelle V, Galaud JP, Thuleau P, Mazars C (2016) Calcium sensors as key hubs in plant responses to biotic and abiotic stresses. Front Plant Sci 7:327CrossRefPubMedPubMedCentralGoogle Scholar
  36. Sasabe M, Takeuchi K, Kamoun S, Ichinose Y, Govers F, Toyoda K, Shiraishi T, Yamada T (2000) Independent pathways leading to apoptotic cell death, oxidative burst and defense gene expression in response to elicitin in tobacco cell suspension culture. FEBS J 267:5005–5013Google Scholar
  37. Stergiopoulos I, de Wit PJ (2009) Fungal effector proteins. Annu Rev Phytopathol 47:233–263CrossRefPubMedGoogle Scholar
  38. Suzaki K (2008) Population structure of Valsa ceratosperma, causal fungus of Valsa canker, in apple and pear orchards. J Gen Plant Pathol 74:128–132CrossRefGoogle Scholar
  39. Tarazona S, García F, Ferrer A, Dopazo J, Conesa A (2012) Noiseq: a rna-seq differential expression method robust for sequencing depth biases. University of Southampton 17:18Google Scholar
  40. Tsegaye Y, Richardson CG, Bravo JE, Mulcahy BJ, Lynch DV, Markham JE, Jaworski JG, Chen M, Cahoon EB, Dunn TM (2007) Arabidopsis mutants lacking long chain base phosphate lyase are fumonisin-sensitive and accumulate trihydroxy-18:1 long chain base phosphate. J Biol Chem 282:28195–28206CrossRefPubMedGoogle Scholar
  41. Tsuda K, Katagiri F, Parker J E, Ellis J G (2010) Comparing signaling mechanisms engaged in patterntriggered and effector-triggered immunity. Curr Opin Plant Biol 13:459-465CrossRefPubMedGoogle Scholar
  42. Vlot AC, Klessig DF, Park SW (2008) Systemic acquired resistance: the elusive signal (s). Curr Opin Plant Biol 11:436–442CrossRefPubMedGoogle Scholar
  43. Wang S, Hu T, Wang Y, Luo Y, Michailides TJ, Cao K (2016) New understanding on infection processes of Valsa canker of apple in China. Eur J Plant Pathol 146:531–540CrossRefGoogle Scholar
  44. Wei J, Huang L, Gao Z, Ke X, Kang Z (2010) Laboratory evaluation methods of apple Valsa canker disease caused by Valsa ceratosperma sensu Kobayashi. Acta Phytopathologica Sinica 40:14–20 (In Chinese)Google Scholar
  45. Ye J, Fang L, Zheng H, Zhang Y, Chen J, Zhang Z, Wang J, Li S, Li R, Bolund L, Wang J (2006) WEGO: a web tool for plotting GO annotations. Nucleic Acids Res 34:W293–W297CrossRefPubMedPubMedCentralGoogle Scholar
  46. Yin Z, Ke X, Kang Z, Huang L (2016) Apple resistance responses against Valsa mali revealed by transcriptomics analyses. Physiol Mol Plant Pathol 93:85–92CrossRefGoogle Scholar
  47. Zhai L, Zhang M, Lv G, Chen X, Jia N, Hong N, Wang G (2014) Biological and molecular characterization of four Botryosphaeria species isolated from pear plants showing stem wart and stem canker in China. Plant Dis 98:716–726CrossRefGoogle Scholar
  48. Zheng Z, Qamar SA, Chen Z, Mengiste T (2006) Arabidopsis WRKY33 transcription factor is required for resistance to necrotrophic fungal pathogens. Plant J 48:592–605CrossRefPubMedGoogle Scholar
  49. Zuo C, Zhang W, Mao J, Jiang X, Ma Z, Su J, Chen B (2017a) Genome wide identification and expression analysis of LysM receptor like kinase in apple. Acta Horticulturae Sinica 44:733–742 (In Chinese)Google Scholar
  50. Zuo C, Zhang W, Chen Z, Chen B, Huang Y (2017b) RNA sequencing reveals that endoplasmic reticulum stress and disruption of membrane integrity underlie dimethyl Trisulfide toxicity against Fusarium oxysporum f. sp. cubense tropical race 4. Front Microbiol 8:1365CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2018

Authors and Affiliations

  • Cunwu Zuo
    • 1
  • Juan Mao
    • 1
  • Zhongjian Chen
    • 2
  • Mingyu Chu
    • 1
  • Hu Duo
    • 1
  • Baihong Chen
    • 1
    Email author
  1. 1.College of HorticultureGansu Agricultural UniversityLanzhouChina
  2. 2.Agro-biological Gene Research CenterGuangdong Academy of Agricultural SciencesGuangzhouChina

Personalised recommendations