Inhibition of pathogenicity of the rice blast fungus by Saccharomyces cerevisiae α-factor

Science

Volumn 276, Issue 5315, 1997, Pages 1116-1119

  Beckerman, Janna L ^a   Naider, Fred ^b   Ebbole, Daniel J ^a  

^a TEXAS A AND M UNIVERSITY   (United States)

^b CITY UNIVERSITY OF NEW YORK   (United States)

Author keywords

[No Author keywords available]

Indexed keywords

MATING HORMONE ALPHA FACTOR; PHEROMONE;

ANTIFUNGAL ACTIVITY; ARTICLE; GENE LOCUS; PATHOGENICITY; PLANT; PRIORITY JOURNAL; RECEPTOR BINDING; SACCHAROMYCES CEREVISIAE;

AMINO ACID SEQUENCE; ASCOMYCOTA; CROSSES, GENETIC; CYCLIC AMP; HORDEUM; MOLECULAR SEQUENCE DATA; ORYZA SATIVA; PEPTIDES; PHEROMONES; PLANT DISEASES; RECEPTORS, MATING FACTOR; RECEPTORS, PEPTIDE; SACCHAROMYCES CEREVISIAE; TRANSCRIPTION FACTORS;

EID: 0030921135     PISSN: 00368075     EISSN: None     Source Type: Journal    
DOI: 10.1126/science.276.5315.1116     Document Type: Article

References (21)

1. B. Valent, Phytopathology 80, 33 (1990).
2. J. E. Hamer, R. J. Howard, F. G. Chumley, B. Valent, Science 239, 288 (1988).
3. R. J. Howard, in Rice Blast Disease, R. S. Zeigler, S. A. Leong, P. S. Teng, Eds. (CAB International, Wallingford, UK, 1994), pp. 3-22; _, M. A. Ferrari, D. H. Roach, N. P. Money, Proc. Natl. Acad. Sci. U.S.A. 88, 11281 (1991).
4. R. J. Howard, in Rice Blast Disease, R. S. Zeigler, S. A. Leong, P. S. Teng, Eds. (CAB International, Wallingford, UK, 1994), pp. 3-22; _, M. A. Ferrari, D. H. Roach, N. P. Money, Proc. Natl. Acad. Sci. U.S.A. 88, 11281 (1991).
5. M. A. Nelson, Trends Genet. 12, 69 (1996).
6. I. Herskowitz, Cell 80, 187 (1995).
7. J. L. Beckerman and D. J. Ebbole, Mol. Plant-Microbe Interact. 9, 450 (1996).
8. The yeast extract from Difco (20 mg/ml) was extracted with benzyl alcohol, and this fraction was extracted with water [J. A. Garibaldi and J. B. Neilands, J. Am. Chem. Soc. 77, 2429 (1955)]. After lyophilization, the residue was dissolved in water and filter sterilized. Difco yeast extract is produced from predominantly diploid cultures and should not contain significant quantities of α-factor. Yeast extract was purchased from Fisher, Sigma, and Difco, but in all cases, it was manufactured by Difco. The different lots obtained were found to have activity toward MAT1-2 but not MAT1-1 strains of M. grisea. The partially purified fraction from yeast extract, unlike α-factor, did not induce mating projection formation or arrest growth in a halo assay when tested against S. cerevisiae strain RC629 (MATa sst1).
9. The yeast extract benzyl alcohol fraction was treated overnight at room temperature with proteinase K at 1 mg/ml or was left untreated as a control. Magnaporthe grisea conidia were mixed directly with these preparations and placed on the hydrophobic side of gel-bond film for appressorium formation assays. In experiments with chymotrypsin-linked beads, the enzyme-linked beads were added (5 mg/ml) to strain CP987 extract or to α-factor and incubated for 12 hours at 25°C. The beads were removed by centrifugation, and the extract was tested for activity. As a control, enzyme-linked beads were heat treated (95°C for 60 min) to destroy protease activity before addition to extract.
10. J. McCullough and I. Herskowitz, J. Bacteriol. 138, 146 (1979); T. Hisatomi, N. Yanagishima, A. Sakurai, H. Kobayashi, Curr. Genet. 13, 25 (1988).
11. J. McCullough and I. Herskowitz, J. Bacteriol. 138, 146 (1979); T. Hisatomi, N. Yanagishima, A. Sakurai, H. Kobayashi, Curr. Genet. 13, 25 (1988).
12. S. K. Raths, F. Naider, J. M. Becker, J. Biol. Chem. 263, 17333 (1988).
13. S. Kang, F. G. Chumley, B. Valent, Genetics 138, 289 (1994).
14. Y.-H. Lee and R. A. Dean, Plant Cell 5, 693 (1993).
15. a-Factor pheromone [C.B. Xue, G.A. Caldwell, J.M. Becker, F. Naider, Biochem. Biophys. Res. Comm. 162, 253 (1989 )] was dissolved in dimethyl sulfoxide to 1 mg/ml and droplets of 1, 5, 10, and 20 μl were placed on the surface of Teflon film (DuPont). The dimethyl sulfoxide was allowed to evaporate for 48 hours, and M. grisea conidia of strains 4091-5-8 and CP987 were placed on the dried a-factor to test for appressorium formation. This same preparation of a-factor was previously shown to elicit growth arrest in a solid agar halo assay and alter cell morphology in suspension with S. cerevisiae strain RC757 [S. Marcus et al., Mol. Cell. Biol. 11, 3603 (1991)].
16. a-Factor pheromone [C.B. Xue, G.A. Caldwell, J.M. Becker, F. Naider, Biochem. Biophys. Res. Comm. 162, 253 (1989 )] was dissolved in dimethyl sulfoxide to 1 mg/ml and droplets of 1, 5, 10, and 20 μl were placed on the surface of Teflon film (DuPont). The dimethyl sulfoxide was allowed to evaporate for 48 hours, and M. grisea conidia of strains 4091-5-8 and CP987 were placed on the dried a-factor to test for appressorium formation. This same preparation of a-factor was previously shown to elicit growth arrest in a solid agar halo assay and alter cell morphology in suspension with S. cerevisiae strain RC757 [S. Marcus et al., Mol. Cell. Biol. 11, 3603 (1991)].
17. Magnaporthe grisea mates and produces perithecia on the mycelial mat formed in standing cultures of oatmeal broth. A culture was coinoculated with strains CP987 and 4091-5-8 and grown until perithecia were visible (about 14 days). Separate flasks were inoculated with strain CP987 or 4091-5-8 and harvested when perithecia formed in the mated control culture. The strain CP987 and 4091-5-8 culture filtrates were harvested, and extracts were prepared as described for yeast extract. A second extract preparation from strain CP987 was used for appressorium assays with strains CP2735 and CP2738. Strains 4136-4-3 and 4091-5-8 produced 92 ± 5% and 8 ± 4% appressoria, respectively, with this second extract preparation. These strain CP987 extracts did not possess activity toward S. cerevisiae strain RC629 in halo assays.
18. 2) (17) with no sequence similarity to yeast pheromone did not protect plants from infection by strain 4091-5-8 at 0.5 mg/ml.
19. E. Loumaye, J. Thorner, K. J. Catt, Science 218, 1323 (1982).
20. Abbreviations for the amino acid residues are: A, Ala; C, Cys; D, Asp; G, Gly; H, His; I, IIe; K, Lys; L, Leu; M, Met; N, Asn; P, Pro; Q, Gln; S, Ser; V, Val; W, Trp; and Y, Tyr.
21. Strains were generously provided by B. Valent of DuPont, Wilmington, DE, and J. Becker of the University of Tennessee, Knoxville. We thank S. McBride for technical assistance and B. Valent and J. E. Hamer for providing research materials and encouragement to work with M. grisea. We also thank J. Becker and D. Cook for helpful discussions. Support for this work was provided by funds from USPHS grant R29GM47977, the Texas Rice Research Foundation, the Texas Advanced Research Program (D.J.E.), and USPHS grant GM22086 (F.N).