Plant Molecular Biology Reporter

, Volume 32, Issue 2, pp 584–595 | Cite as

Identification of Late Blight Resistance-Related Metabolites and Genes in Potato through Nontargeted Metabolomics

  • Doddaraju Pushpa
  • Kalenahalli N. Yogendra
  • Raghavendra Gunnaiah
  • Ajjamada C. Kushalappa
  • Agnes Murphy
Original Paper

Abstract

Late blight of potato (Solanum tuberosum) caused by Phytophthora infestans significantly reduces the productivity of potato around the world. Resistance to late blight in potato is either qualitative or quantitative. Qualitative resistance governed by race-specific single R genes is well characterized and gives complete resistance, but is not durable. Quantitative resistance governed by polygenes gives partial resistance, but is durable in nature. However, the quantitative resistance mechanisms are poorly studied and are not efficiently exploited in potato breeding. A nontargeted metabolic profiling of resistant (F06037) and susceptible (Shepody) potato cultivars, using high-resolution liquid chromatography–mass spectrometry, was applied to elucidate the quantitative resistance mechanisms against P. infestans (US-8 genotype). The hydroxycinnamic acid amides (HCAAs) of the shunt phenylpropanoid pathway were highly induced following pathogen inoculation in F06037. In parallel, the transcript abundances of genes that catalyze the biosynthesis of these metabolites, such as 4-coumarate:CoA ligase, tyrosine decarboxylase, ornithine decarboxylase, tyramine hydroxycinnamoyl transferase, and putrescine hydroxycinnamoyl transferase, were also higher in the resistant genotype. Sequencing of the coding genes of these enzymes revealed single-nucleotide polymorphisms between resistant and susceptible genotypes, and the amino acid changes caused missense mutations altering protein functions. HCAAs deposited at host cell walls inhibit pathogen colonization, thus reducing lesion expansion. In addition, these also act as phytoalexins, leading to the reduced biomass of the pathogen. Following validation, the HCAAs can be used as biomarker metabolites for late blight resistance. The putative candidate genes can be either used to develop allele-specific markers for marker-assisted breeding programs or suitably stacked into elite cultivars through cisgenic approaches, following validation.

Keywords

Metabolomics Quantitative resistance Phytophthora infestans Polygenic resistance Single-nucleotide polymorphism Potato 

Supplementary material

11105_2013_665_Fig6_ESM.jpg (104 kb)
Fig. S1

MS/MS spectra and in silico fragmentation of metabolites detected in potato cultivars with contrasting levels of late blight resistance following P. infestans or mock inoculation. a) p-Coumaroyltyramine, b) sinapoyltyramine. (JPEG 103 kb)

11105_2013_665_MOESM1_ESM.tif (259 kb)
High resolution image(TIFF 259 kb)
11105_2013_665_Fig7_ESM.jpg (57 kb)
Fig. S2

Resistant related metabolites are clustered in to two groups mainly RRC and RRI. Their chemical groups and fold change are indicated as node shape and color. RRC is resistant related constitutive metabolites, RRI is resistant related induced metabolites and NI and NS is not identified in our study and non-significant at (P < 0.05). (JPEG 57 kb)

11105_2013_665_MOESM2_ESM.tif (15.2 mb)
High resolution image(TIFF 15516 kb)
11105_2013_665_MOESM3_ESM.xls (40 kb)
Table S1Late blight resistance related constitutive metabolites identified in potato cultivars following P. infestans or mock inoculation. (XLS 40 kb)
11105_2013_665_MOESM4_ESM.xls (36 kb)
Table S2Late blight resistance related induced metabolites identified in potato cultivars following P. infestans or mock inoculation. (XLS 36 kb)

References

  1. Abu-Nada KAC (2010) Metabolic profiling to phenotype potato cultivars varying in horizontal resistance to leaf infection by Phytophthora infestans. Am J Plant Sci Biotechnol 4(2):55–64Google Scholar
  2. Agati G, Azzarello E, Pollastri S, Tattini M (2012) Flavonoids as antioxidants in plants: location and functional significance. Plant Sci 196:67–76Google Scholar
  3. Andrivon D, Corbière R, Lucas JM, Pasco C, Gravoueille JM, Pellé R, Dantec JP, Ellissèche D (2003) Resistance to late blight and soft rot in six potato progenies and glycoalkaloid contents in the tubers. Am J Potato Res 80(2):125–134CrossRefGoogle Scholar
  4. Arman M (2011) LC-ESI-MS characterisation of phytoalexins induced in chickpea and pea tissues in response to a biotic elicitor of Hypnea musciformis (red algae). Nat Prod Res 25(14):1352–1360PubMedCrossRefGoogle Scholar
  5. Ashkenazy H, Erez E, Martz E, Pupko T, Ben-Tal N (2010) ConSurf 2010: calculating evolutionary conservation in sequence and structure of proteins and nucleic acids. Nucleic Acids Res 38(suppl 2):W529–W533PubMedCentralPubMedCrossRefGoogle Scholar
  6. Bassard JE, Ullmann P, Bernier F, Werck-Reichhart D (2010) Phenolamides: bridging polyamines to the phenolic metabolism. Phytochemistry 71(16):1808–1824PubMedCrossRefGoogle Scholar
  7. Bernards MA, Lopez ML, Zajicek J, Lewis NG (1995) Hydroxycinnamic acid-derived polymers constitute the polyaromatic domain of suberin. J Biol Chem 270(13):7382–7386PubMedCrossRefGoogle Scholar
  8. Birch PRJ, Bryan G, Fenton B, Gilroy EM, Hein I, Jones JT, Prashar A, Taylor MA, Torrance L, Toth IK (2012) Crops that feed the world 8: Potato: are the trends of increased global production sustainable? Food Security 4:477–508Google Scholar
  9. Blount JW, Dixon RA, Paiva NL (1992) Stress responses in alfalfa (Medicago sativa L.) XVI. Antifungal activity of medicarpin and its biosynthetic precursors; implications for the genetic manipulation of stress metabolites. Physiol Mol Plant Pathol 41(5):333–349CrossRefGoogle Scholar
  10. Bollina V, Kumaraswamy GK, Kushalappa AC, Choo TM, Dion Y, Rioux S, Faubert D, Hamzehzarghani H (2010) Mass spectrometry–based metabolomics application to identify quantitative resistance–related metabolites in barley against Fusarium head blight. Mol Plant Pathol 11(6):769–782PubMedGoogle Scholar
  11. Bryant D, Cummins I, Dixon DP, Edwards R (2006) Cloning and characterization of a theta class glutathione transferase from the potato pathogen Phytophthora infestans. Phytochemistry 67(14):1427–1434PubMedCrossRefGoogle Scholar
  12. Cho K, Kim Y, Wi SJ, Seo JB, Kwon J, Chung JH, Park KY, Nam MH (2012) Nontargeted metabolite profiling in compatible pathogen-inoculated tobacco (Nicotiana tabacum L. cv. Wisconsin 38) using UPLC-Q-TOF/MS. J Agric Food Chem 60(44):11015–11028PubMedCrossRefGoogle Scholar
  13. Clarke D (1982) The accumulation of cinnamic acid amides in the cell walls of potato tissue as an early response to fungal attack. In: Wood RKS (ed) Active defense mechanisms in plants. Plenum, London, 321 ppGoogle Scholar
  14. Cowley T, Walters D (2002) Polyamine metabolism in barley reacting hypersensitively to the powdery mildew fungus Blumeria graminis f. sp. hordei. Plant Cell Environ 25(3):461–468CrossRefGoogle Scholar
  15. El-Bebany AF, Adam LR, Daayf F (2013) Differential accumulation of phenolic compounds in potato in response to weakly and highly aggressive isolates of Verticillium dahliae. Can J Plant Pathol 35(2):232–240CrossRefGoogle Scholar
  16. Fewell AM, Roddick JG (1997) Potato glycoalkaloid impairment of fungal development. Mycol Res 101(5):597–603CrossRefGoogle Scholar
  17. Fiehn O (2002) Metabolomics—the link between genotypes and phenotypes. Plant Mol Biol 48(1–2):155–171PubMedCrossRefGoogle Scholar
  18. Fleurence J, Negrel J (1989) Partial purification of tyramine feruloyl transferase from TMV inoculated tobacco leaves. Phytochemistry 28(3):733–736CrossRefGoogle Scholar
  19. Friedman M (2006) Potato glycoalkaloids and metabolites: roles in the plant and in the diet. J Agric Food Chem 54(23):8655–8681PubMedCrossRefGoogle Scholar
  20. Fry W (2008) Phytophthora infestans: the plant (and R gene) destroyer. Mol Plant Pathol 9(3):385–402PubMedCrossRefGoogle Scholar
  21. Fry WE, Goodwin SB (1997) Resurgence of the Irish potato famine fungus. BioScience 47:363–371Google Scholar
  22. Gebhardt C (2012) Bridging the gap between genome analysis and precision breeding in potato. Trends Genet 29:248–256Google Scholar
  23. González-Coloma A, López-Balboa C, Santana O, Reina M, Fraga BM (2011) Triterpene-based plant defenses. Phytochem Rev 10(2):245–260CrossRefGoogle Scholar
  24. Gunnaiah R, Kushalappa AC, Duggavathi R, Fox S, Somers DJ (2012) Integrated metabolo-proteomic approach to decipher the mechanisms by which wheat QTL (Fhb1) contributes to resistance against Fusarium graminearum. PLoS ONE 7(7):e40695PubMedCentralPubMedCrossRefGoogle Scholar
  25. Hagel JM, Facchini PJ (2005) Elevated tyrosine decarboxylase and tyramine hydroxycinnamoyltransferase levels increase wound-induced tyramine-derived hydroxycinnamic acid amide accumulation in transgenic tobacco leaves. Planta 221(6):904–914PubMedCrossRefGoogle Scholar
  26. Haverkort A, Struik P, Visser RGF, Jacobsen E (2009) Applied biotechnology to combat late blight in potato caused by Phytophthora infestans. Potato Res 52(3):249–264CrossRefGoogle Scholar
  27. Katagiri Y, IBRAHIM RK, TAHARA S (2000) HPLC analysis of white lupin isoflavonoids. Biosci Biotechnol Biochem 64(6):1118–1125PubMedCrossRefGoogle Scholar
  28. Kaur H, Heinzel N, Schöttner M, Baldwin IT, Gális I (2010) R2R3-NaMYB8 regulates the accumulation of phenylpropanoid-polyamine conjugates, which are essential for local and systemic defense against insect herbivores in Nicotiana attenuata. Plant Physiol 152(3):1731–1747PubMedCentralPubMedCrossRefGoogle Scholar
  29. Keller H, Hohlfeld H, Wray V, Hahlbrock K, Scheel D, Strack D (1996) Changes in the accumulation of soluble and cell wall-bound phenolics in elicitor-treated cell suspension cultures and fungus-infected leaves of Solanum tuberosum. Phytochemistry 42(2):389–396CrossRefGoogle Scholar
  30. Kolattukudy P (1981) Structure, biosynthesis, and biodegradation of cutin and suberin. Annu Rev Plant Physiol 32(1):539–567CrossRefGoogle Scholar
  31. Kolattukudy P (1984) Biochemistry and function of cutin and suberin. Can J Bot 62(12):2918–2933CrossRefGoogle Scholar
  32. Kou Y, Wang S (2010) Broad-spectrum and durability: understanding of quantitative disease resistance. Curr Opin Plant Biol 13(2):181–185PubMedCrossRefGoogle Scholar
  33. Kumar P, Henikoff S, Ng PC (2009) Predicting the effects of coding non-synonymous variants on protein function using the SIFT algorithm. Nat Protocol 4(7):1073–1081CrossRefGoogle Scholar
  34. Kumaraswamy GK, Bollina V, Kushalappa AC, Choo TM, Dion Y, Rioux S, Mamer O, Faubert D (2011) Metabolomics technology to phenotype resistance in barley against Gibberella zeae. Eur J Plant Pathol 130(1):29–43CrossRefGoogle Scholar
  35. Kushalappa AC, Gunnaiah R (2013) Metabolo-proteomics to discover plant biotic stress resistance genes. Trends Plant Sci 18:522–531PubMedCrossRefGoogle Scholar
  36. Langebartels C, Kerner K, Leonardi S, Schraudner M, Trost M, Heller W, Sandermann H (1991) Biochemical plant responses to ozone I. Differential induction of polyamine and ethylene biosynthesis in tobacco. Plant Physiol 95(3):882–889PubMedCentralPubMedCrossRefGoogle Scholar
  37. Menden B, Kohlhoff M, Moerschbacher BM (2007) Wheat cells accumulate a syringyl-rich lignin during the hypersensitive resistance response. Phytochemistry 68(4):513–520PubMedCrossRefGoogle Scholar
  38. Mustafa NR, Verpoorte R (2007) Phenolic compounds in Catharanthus roseus. Phytochem Rev 6(2):243–258CrossRefGoogle Scholar
  39. Naoumkina MA, Zhao Q, Gallego-giraldo L, Dai X, Zhao PX, Dixon RA (2010) Genome-wide analysis of phenylpropanoid defence pathways. Mol Plant Pathol 11(6):829–846Google Scholar
  40. Negrel J, Pollet B, Lapierre C (1996) Ether-linked ferulic acid amides in natural and wound periderms of potato tuber. Phytochemistry 43(6):1195–1199CrossRefGoogle Scholar
  41. Ng PC, Henikoff S (2003) SIFT: predicting amino acid changes that affect protein function. Nucleic Acids Res 31(13):3812–3814PubMedCentralPubMedCrossRefGoogle Scholar
  42. Nick A, Wright AD, Rali T, Sticher O (1995) Antibacterial triterpenoids from Dillenia papuana and their structure–activity relationships. Phytochemistry 40(6):1691–1695PubMedCrossRefGoogle Scholar
  43. Nicot N, Hausman J-F, Hoffmann L, Evers D (2005) Housekeeping gene selection for real-time RT-PCR normalization in potato during biotic and abiotic stress. J Exp Bot 56(421):2907–2914PubMedCrossRefGoogle Scholar
  44. Osbourn AE (1996) Preformed antimicrobial compounds and plant defense against fungal attack. Plant Cell 8(10):1821PubMedCentralPubMedGoogle Scholar
  45. Picman AK (1986) Biological activities of sesquiterpene lactones. Biochem Syst Ecol 14(3):255–281CrossRefGoogle Scholar
  46. Pluskal T, Castillo S, Villar-Briones A, Orešič M (2010) MZmine 2: modular framework for processing, visualizing, and analyzing mass spectrometry-based molecular profile data. BMC Bioinforma 11(1):395CrossRefGoogle Scholar
  47. Ponts N, Pinson-Gadais L, Boutigny A-L, Barreau C, Richard-Forget F (2011) Cinnamic-derived acids significantly affect Fusarium graminearum growth and in vitro synthesis of type B trichothecenes. Phytopathology 101(8):929–934PubMedCrossRefGoogle Scholar
  48. Prost I, Dhondt S, Rothe G, Vicente J, Rodriguez MJ, Kift N, Carbonne F, Griffiths G, Esquerré-Tugayé M-T, Rosahl S (2005) Evaluation of the antimicrobial activities of plant oxylipins supports their involvement in defense against pathogens. Plant Physiol 139(4):1902–1913PubMedCentralPubMedCrossRefGoogle Scholar
  49. Ranathunge K, Schreiber L, Franke R (2011) Suberin research in the genomics era—new interest for an old polymer. Plant Sci 180(3):399–413PubMedCrossRefGoogle Scholar
  50. Reina-Pinto JJ, Yephremov A (2009) Surface lipids and plant defenses. Plant Physiol Biochem 47(6):540–549PubMedCrossRefGoogle Scholar
  51. Riley RG, Kolattukudy PE (1975) Evidence for covalently attached p-coumaric acid and ferulic acid in cutins and suberins. Plant Physiol 56(5):650–654PubMedCentralPubMedCrossRefGoogle Scholar
  52. Roy A, Kucukural A, Zhang Y (2010) I-TASSER: a unified platform for automated protein structure and function prediction. Nat Protocol 5(4):725–738CrossRefGoogle Scholar
  53. Schauer N, Fernie AR (2006) Plant metabolomics: towards biological function and mechanism. Trend Plant Sci 11(10):508–516CrossRefGoogle Scholar
  54. Secor GA, Gudmestad NC (1999) Managing fungal diseases of potato. Can J Plant Pathol 21(3):213–221CrossRefGoogle Scholar
  55. Shannon P, Markiel A, Ozier O, Baliga NS, Wang JT, Ramage D, Amin N, Schwikowski B, Ideker T (2003) Cytoscape: a software environment for integrated models of biomolecular interaction networks. Genome Res 13(11):2498–2504PubMedCrossRefGoogle Scholar
  56. Simko I (2002) Comparative analysis of quantitative trait loci for foliage resistance to Phytophthora infestans in tuber-bearing Solanum species. Am J Potato Res 79(2):125–132CrossRefGoogle Scholar
  57. Stewart H, Bradshaw J, Pande B (2003) The effect of the presence of R-genes for resistance to late blight (Phytophthora infestans) of potato (Solanum tuberosum) on the underlying level of field resistance. Plant Pathol 52(2):193–198CrossRefGoogle Scholar
  58. Thomas R, Fang X, Ranathunge K, Anderson TR, Peterson CA, Bernards MA (2007) Soybean root suberin: anatomical distribution, chemical composition, and relationship to partial resistance to Phytophthora sojae. Plant Physiol 144(1):299–311PubMedCentralPubMedCrossRefGoogle Scholar
  59. Thompson JD, Gibson TJ, Plewniak F, Jeanmougin F, Higgins DG (1997) The CLUSTAL_X windows interface: flexible strategies for multiple sequence alignment aided by quality analysis tools. Nucleic Acid Res 25(24):4876–4882PubMedCrossRefGoogle Scholar
  60. Vanholme R, Demedts B, Morreel K, Ralph J, Boerjan W (2010) Lignin biosynthesis and structure. Plant Physiol 153(3):895–905PubMedCentralPubMedCrossRefGoogle Scholar
  61. Walters DR (2003) Polyamines and plant disease. Phytochemistry 64(1):97–107PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2013

Authors and Affiliations

  • Doddaraju Pushpa
    • 1
  • Kalenahalli N. Yogendra
    • 1
  • Raghavendra Gunnaiah
    • 1
  • Ajjamada C. Kushalappa
    • 1
  • Agnes Murphy
    • 2
  1. 1.Plant Science DepartmentMcGill UniversitySainte-Anne-de-BellevueCanada
  2. 2.Agriculture and Agri-Food CanadaFrederictonCanada

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