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Identification and chromosomal locations of a family of cytochrome P-450 genes for pisatin detoxification in the fungus Nectrla haematococca

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Summary

The ability to detoxify the phytoalexin, pisatin, an antimicrobial compound produced by pea (Pisum sativum L.), is one requirement for pathogenicity of the fungus Nectria haematococca on this plant. Detoxification is mediated by a cytochrome P-450, pisatin demethylase, encoded by any one of six Pda genes, which differ with respect to the inducibility and level of pisatin demethylase activity they confer, and which are associated with different levels of virulence on pea. A previously cloned Pda gene (PdaT9) was used in this study to characterize further the known genes and to identify additional members of the Pda family in this fungus by Southern analysis. DNA from all isolates which demethylate pisatin (Pda+ isolates) hybridized to PdaT9, while only one Pda isolate possessed DNA homologous to the probe. Hybridization intensity and, in some cases, restriction fragment size, were correlated with enzyme inducibility. XhoI/BamHI restricted DNA from reference strains with a single active Pda allele had only one fragment with homology to PdaT9; no homology attributable to alleles associated with the Pda phenotype was found. Homology to this probe was also limited to one or two restriction fragments in most of the 31 field isolates examined. Some unusual progeny from laboratory crosses that failed to inherit demethylase activity also lost the single restriction fragment homologous to PdaT9. At the chromosome level, N. haematococca is highly variable, each isolate having a unique electrophoretic karyotype. In most instances, PdaT9 hybridized to one or two chromosomes containing 1.6–2 million bases of DNA, while many Pda- isolates lacked chromosomes in this size class. The results from this study of the Pda family support the hypothesis that deletion of large amounts of genomic DNA is one mechanism that reduces the frequency of Pda genes in N. haematococca, while simultaneously increasing its karyotypic variation.

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References

  1. Atchison M, Adesnik M (1983) A cytochrome P-450 multigene family: characterization of a gene activated by phenobarbital administration. J Biol Chem 258:11285–11295

  2. Brody H, Carbon J (1989) Electrophoretic karyotype of Aspergillus nidulans. Proc Natl Acad Sci USA 86:6260–6263

  3. Cantor CR, Smith CL, Mathews MK (1988) Pulsed-field gel electrophoresis of very large DNA molecules. Annu Rev Biophys Biophys Chem 17:287–304

  4. Carle GF, Olson M (1985) An electrophoretic karyotype for yeast. Proc Natl Acad Sci USA 82:3756–3760

  5. Chu G, Vollrath D, Davis R (1986) Separation of large DNA molecules by contour clamped homogeneous electric fields. Science 234:1582–1585

  6. Fan J-B, Yuji C, Smith CL, Niwa O, Yanagida M, Cantor CR (1989) Construction of a NotI restriction map of the fission yeast Schizosaccharomyces pombe genome. Nucleic Acids Res 17:2801–2818

  7. Geck P, Nasz I (1983) Concentrated, digestible DNA after hydroxylapatite chromatography with cetylpyridinium bromide precipitation. Anal Biochem 135:264–268

  8. Gibson WC, Borst P (1986) Size fractionation of the small chromosomes of Trypanozoon and Nannomonas trypanosomes by pulsed field gradient gel electrophoresis. Mot Biochem Parasitol 18:127–140

  9. Johnston JR, Mortimer RK (1986) Electrophoretic karyotyping of laboratory and commercial strains of Saccharomyces and other yeasts. Int J System Bacteriol 36: 569–572

  10. Jones RN, Rees H (1982) B chromosomes. Academic Press, NY

  11. Kinscherf TG, Leong SA (1988) Molecular analysis of the karyotype of Ustilago maydis. Chromosoma 96:427–433

  12. Kistler HC, VanEtten HD (1984) Three non-allelic genes for pisatin demethylation in the fungus Nectria haematococca. J Gen Microbiol 130:2595–2603

  13. Lucy MC, Matthews PS, VanEtten HD (1988) Metabolic detoxification of the phytoalexins maackiain and medicarpin in Nectria haematococca field isolates: relationship to virulence on chickpea. Physiol Plant Pathol 33:187–199

  14. Mackintosh SF, Matthews DE, VanEtten HD (1989) Two additional genes for pisatin demethylation and their relationship to the pathogenicity of Nectria haematococca on pea. Mot Plant-Microbe Interact 2:354–362

  15. Magee BB, Magee PT (1987) Electrophoretic karyotypes and chromosome numbers in Candida species. J Gen Microbiol 133:425–430

  16. Magee BB, Koltin Y, Gorman JA, Magee PT (1988) Assignment of cloned genes to the seven electrophoretically separated Candida albicans chromosomes. Mol Cell Biol 8:4721–4726

  17. Maniotis J (1979) Polyploidy in fungi. In: Lewis WH (ed) Polyploidy: biological significance. Plenum Press, NY, pp 163–192

  18. Matthews DE, VanEtten HD (1983) Detoxification of the phytoalexin pisatin by a fungal cytochrome P-450. Arch Biochem Biophys 224:494–505

  19. Nebert DW, Nelson DR, Feyereisen R (1989) Evolution of the cytochrome P450 genes. Xenobiotica 19:1149–1160

  20. Nelson DR, Strobel HW (1987) Evolution of cytochrome P-450 proteins. Mot Biol Evol 4: 572–593

  21. Orbach M, Vollrath D, Davis RW, Yanofsky C (1988) An electrophoretic karyotype of Neurospora crassa. Mot Cell Biol 8:1469–1473

  22. Puhalla J (1981) Genetic considerations of the genus Fusarium. In: Nelson PE, Toussoun TA, Cook RJ (eds) Fusarium: diseases, biology and taxonomy. Pennsylvania State University Press, University Park, Pennsylvania, pp 291–305

  23. Schäfer W, Straney D, Ciuffetti L, VanEtten HD, Yoder OC (1989) One enzyme makes a fungal pathogen, but not a saprophyte, virulent on a new host plant. Science 246:247–249

  24. Schwartz C, Cantor CR (1984) Separation of yeast-chromosome sized DNAs by pulsed field gradient gel electrophoresis. Cell 37:67–75

  25. Smith DA, Banks SW (1986) Biosynthesis, elicitation and biological activity of isoflavonoid phytoalexins. Phytochemistry 25:979–995

  26. Tegtmeier KJ, VanEtten HD (1982) The role of pisatin tolerance and degradation in the virulence of Nectria haematococca on peas: a genetic analysis. Phytopathology 72:608–612

  27. Tolmsoff WJ (1983) Heteroploidy as a mechanism of variability among fungi. Annu Rev Phytopathol 21:317–340

  28. Van der Ploeg LHT, Smits M, Ponnudurai T, Vermeulen A, Meuwissen JHETh, Langley G (1985) Chromosome-sized DNA molecules of Plasmodium falciparum. Science 229: 658–661

  29. VanEtten HD, Matthews PS (1984) Naturally occurring variation in the inducibility of pisatin demethylating activity in Nectria haematococca mating population VI. Physiol Plant Pathol 25:149–160

  30. VanEtten HD, Stein JI (1978) Differential response of Fusarium solani isolates to pisatin and phaseollin. Phytopathology 68:1276–1283

  31. VanEtten HD, Pueppke SG, Kelsey TC (1975) 3,6a dihydroxy-8,9,methylenedioxypterocarpan as a metabolite of pisatin produced by Fusarium solani f. sp. pisi. Phytochemistry 14:1103–1105

  32. VanEtten HD, Matthews PS, Tegtmeier KJ, Dietert MF, Stein J (1980) The association of pisatin tolerance and demethylation with virulence on pea in Nectria haematococca. Physiol Plant Pathol 16:257–268

  33. VanEtten HD, Matthews DE, Matthews PS (1989a) Phytoalexin detoxification: importance for pathogenicity and practical implications. Annu Rev Phytopathol 27:143–164

  34. VanEtten HD, Matthews DE, Matthews P, Miao V, Maloney A, Straney D (1989b) A family of genes for phytoalexin detoxification in the plant pathogen Nectria haematococca. In: Lugtenberg B (ed) Signal Molecules in Plants and Plant-Microbe Interactions, NATO ASI series, Vol H36: Springer-Verlag, Berlin, pp 219–228

  35. Vollrath D, Davis RW (1987) Resolution of DNA molecules greater than 5 megabases by contour clamped homogeneous electric fields. Nucleic Acids Res 15:7865–7876

  36. Vollrath D, Davis RW, Connelly C, Hieter P (1988) Physical mapping of large DNA by chromosome fragmentation. Proc Natl Acad Sci USA 85:6027–6031

  37. Weltring KM, Turgeon BG, Yoder OC, VanEtten HD (1988) Isolation of a phytoalexin-detoxification gene from the plant pathogenic fungus Nectria haematococca by detecting its expression in Aspergillus nidulans. Gene 68:335–344

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Communicated by C.A.M. van den Hondel

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Miao, V.P.W., Matthews, D.E. & VanEtten, H.D. Identification and chromosomal locations of a family of cytochrome P-450 genes for pisatin detoxification in the fungus Nectrla haematococca . Mol Gen Genet 226, 214–223 (1991). https://doi.org/10.1007/BF00273606

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Key words

  • Phytoalexin
  • Pisatin demethylase
  • Cytochrome P-450
  • Plant pathogen
  • Electrophoretic karyotype