Lipids

, Volume 35, Issue 3, pp 243–247 | Cite as

Biochemical modifications and transcriptional alterations attendant to sterol feeding in Phytophthora parasitica

  • W. David Dotson
  • Shirley R. Tove
  • Leo W. Parks
Article

Abstract

Phytophthora species are eukaryotic sterol auxotrophs that possess the ability to grow, albeit poorly, in the complete absence of sterols. Growth of Phytophthora is often improved substantially when an exogenous source of sterol is provided. Additionally, sterols may be required for sexual and asexual sporulation in Phytophthora. Our research has been focused on identifying and characterizing the immediate physiological effects following sterol addition to cultures of P. parasitica. Through gas chromatographic analysis of extracts from P. parasitica cultures that were fed various sterols, we have ob-tained evidence for sterol C5 desaturase and Δ7 reductase activities in this organism. Zoo blots were probed with DNA sequences encoding these enzymes, from Saccharomyces cerevisiae and Arabidopsis thaliana. Hybridization of a S. cerevisiae ERG3 probe to P. parasitica DNA was observed, implicating sequence similarity between the sterol C5 desaturase encoding genes. Differential display experiments, using RNA from P. parasitica, have demonstrated a pattern of altered gene expression between cultures grown in the presence and absence of sitosterol. Characterization of sterol-related metabolic effects and sterol functions in Phytophthora should lead to improved measures for control of this important group of plant pathogens.

Abrreviations

Brassicasterol

(22E)-ergosta-5,22-dien-3β-ol

DDRT-PCR

differential display reverse transcriptase polymerase chain reaction

ergosterol

(22E)-ergosta-5,7,22-trien-3β-ol

GC

gas chromatography

HT11VN

modified oligo-dT anchored primers

lamosterol

lanosta-8,24-dient-3β-ol

PB

Phytophthora basal medium

RRT

Telative retention time

shosterol

stigmast-5-en-3β-ol

TLC

thin-layer chromatography

zymosterol

5α-cholesta-8,24-dien-3β-ol

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References

  1. 1.
    Hendrix, J.W. (1970) Sterols in Growth and Reproduction of Fungi, Annu. Rev. Phytopathol. 8, 111–130.CrossRefGoogle Scholar
  2. 2.
    Jee, H.J., Tang, C.S., and Ko, W.H. (1997) Stimulation of Sexual Reproduction in Phytophthora cactorum by Phospholipids Is Not Due to Sterol Contamination, Microbiology 143, 1631–1638.CrossRefGoogle Scholar
  3. 3.
    Mikes, V., Milat, M.L., Ponchet, M., Ricci, P., and Blein, J.P. (1997) The Fungal Elicitor Cryptogein Is a Sterol Carrier Protein, FEBS Lett. 416, 190–192.PubMedCrossRefGoogle Scholar
  4. 4.
    Choi, D., Ward, B.L., and Bostock, R.M. (1992) Differential Induction and Suppression of Potato 3-Hydroxy-3-methylglutaryl-coenzyme A Reductase Genes in Response to Phytophthora infestans and Its Elicitor Arachidonic Acid, Plant Cell 4, 1333–1344.PubMedCrossRefGoogle Scholar
  5. 5.
    Goffeau, A., Barrell, B.G., Bussey, H., Davis, R.W., Dujon, B., Feldman, H., Galibert, F., Hoheisel, J.D., Jacq, C., Johnston, M., Louis, E.J., Mewes, H.W., Murakami, Y., Philippsen, P., Tettelin, H., and Oliver, S.G. (1996) Life with 6000 Genes, Science 274, 546–567.PubMedCrossRefGoogle Scholar
  6. 6.
    Liang, P., and Pardee, A.B. (1992) Differential Display of Eukaryotic Messenger RNA by Means of the Polymerase Chain Reaction, Science 257, 967–971.PubMedCrossRefGoogle Scholar
  7. 7.
    Liang, P., and Pardee, A.B. (1993) Distribution and Cloning of Eukaryotic mRNAs by Means of Differential Display: Refinements and Optimization, Nucleic Acids Res. 21, 3269–3275.PubMedGoogle Scholar
  8. 8.
    Smith, S.J., and Parks, L.W. (1993) The ERG3 Gene in Saccharomyces cerevisiae Is Required for the Utilization of Respiratory Substrates and in Heme-Deficient Cells, Yeast 9, 1177–1187.PubMedCrossRefGoogle Scholar
  9. 9.
    Smith, S.J., Crowley, J.H., and Parks, L.W. (1993) Transcriptional Regulation by Ergosterol in the Yeast Saccharomyces cerevisiae, Mol. Cell. Biol. 16, 5427–5432.Google Scholar
  10. 10.
    Gonzales, R.A. (1981) The Development of Phytophthora cactorum as a Model System for the Study of Sterol Metabolism, Ph.D. Thesis, Oregon State University, Corvallis, pp. 23–30.Google Scholar
  11. 11.
    Parks, L.W., Bottema, C.D.K., Rodriguez, R.J., and Lewis, T.A. (1985) Yeast Sterols: Yeast Mutants as Tools for the Study of Sterol Metabolism, Methods Enzymol. 111, 333–346.PubMedCrossRefGoogle Scholar
  12. 12.
    Goodwin, S.B., Drenth, A., and Fry, W.E. (1992) Cloning and Genetic Analyses of Two Highly Polymorphic, Moderately Repetitive Nuclear DNAs from Phytophthora infestans, Curr. Genet. 22, 107–115.PubMedCrossRefGoogle Scholar
  13. 13.
    Collart, M.A., and Oliviero, S. (1993) Preparation of Yeast RNA, Curr. Protoc. Mol. Biol. 2, 13.12.1–13.12.5.Google Scholar
  14. 14.
    Sampbrook, J., Fritsch, E.R., and Maniatis, T. (1989) Molecular Cloning: A Laboratory Manual, 2nd edn., Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York, pp. 9.31–9.62.Google Scholar
  15. 15.
    Leak, F.W. (1997) The Phenotypic Diversity of the upc 2 Mutation in Saccharomyces cerevisiae, M.S. Thesis, North Carolina State University, Raleigh, pp. 33–34.Google Scholar
  16. 16.
    Lecain, E., Chenivesse, X., Spagnoli, R., and Pompon, D. (1996) Cloning by Metabolic Interference in Yeast and Enzymatic Characterization of Arabidopsis thaliama Sterol Delta 7-Reductase, J. Biol. Chem. 271, 10866–10873.PubMedCrossRefGoogle Scholar
  17. 17.
    Goad, J.L., and Toshihira, A. (1997) Gas-Liquid Chromatography of Sterols, in Analysis of Sterols, pp. 123–131. Chapman and Hall, New York.Google Scholar
  18. 18.
    Lamparczyk, H. (1992) Sterols, in Analysis and Characterization of Steroids, pp. 119–137, CRC Press, Boca Raton.Google Scholar
  19. 19.
    Knights, B.A., and Elliot, C.G. (1976) Metabolism of Δ7 and Δ5,7 by Phytophthora cactorum, Biochim. Biophys. Acta 441, 341–346.PubMedGoogle Scholar
  20. 20.
    Holmer, L., Pezhman, A., and Worman, H.J. (1998) The Human Lamin B Receptor/Sterol Reductase Multigene Family, Genomics 15, 469–476.CrossRefGoogle Scholar
  21. 21.
    Dudler, R. (1990) The Single-Copy Actin Gene of Phytophthora megasperma Encodes a Protein Considerably Diverged from Any Other Known Actin, Plant Mol. Biol. 14, 415–422.PubMedCrossRefGoogle Scholar
  22. 22.
    Unkles, S.E., Moon, R.P., Hawwkins, A.R., Duncan, J.M., and Kinghorn, J.R. (1991) Actin in the Oomycetous Fungus Phytophthora infestans Is the Product of Several Genes, Gene 100, 105–112.PubMedCrossRefGoogle Scholar

Copyright information

© AOCS Press 2000

Authors and Affiliations

  • W. David Dotson
    • 1
  • Shirley R. Tove
    • 1
  • Leo W. Parks
    • 1
  1. 1.Department of MicrobiologyNorth Carolina State UniversityRaleigh

Personalised recommendations