1887

Abstract

This paper describes (i) the expression profile of the methionine synthase gene () in the human pathogenic fungus and (ii) the phenotypes of a mutant. In contrast to the gene, which showed no significant change in expression in any environmental condition tested, the gene showed a substantial induction in response to methionine and a dramatic transcriptional induction in response to homocysteine. Like a mutant, the mutant was a methionine auxotroph. However, relative to a mutant, the mutant grew very slowly and was less heat-shock resistant. In contrast to a mutant, the mutant lost viability when starved of methionine, and it was deficient in capsule formation. Like a mutant, the mutant was avirulent. In contrast to a mutant, the mutant was hypersensitive to fluconazole and to the calcineurin inhibitors FK506 and cyclosporin A. A synergistic fungicidal effect was also found between each of these drugs and . The phenotypic differences between the and mutants may be due to the accumulation in mutants of homocysteine, a toxic metabolic intermediate that inhibits sterol biosynthesis.

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2004-09-01
2021-02-28
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References

  1. Alspaugh J. A., Perfect J. R., Heitman J. 1997; Cryptococcus neoformans mating and virulence are regulated by the G-protein α subunit GPA1 and cAMP. Genes Dev 11:3206–3217 [CrossRef]
    [Google Scholar]
  2. Alspaugh J. A., Pukkila-Worley R., Harashima T., Cavallo L. M., Funnell D., Cox G. M., Perfect J. R., Kronstad J. W., Heitman J. 2002; Adenylyl cyclase functions downstream of the Gα protein Gpa1 and controls mating and pathogenicity of Cryptococcus neoformans. Eukaryot Cell 1:75–84 [CrossRef]
    [Google Scholar]
  3. Banerjee R. V., Matthews R. G. 1990; Cobalamin-dependent methionine synthase. FASEB J 4:1450–1459
    [Google Scholar]
  4. Barrett D. 2002; From natural products to clinically useful antifungals. Biochim Biophys Acta 1587:224–233 [CrossRef]
    [Google Scholar]
  5. Boguslawski G., Stetler D. A. 1979; Aspects of physiology of Histoplasma capsulatum. Mycopathologia 67:17–24 [CrossRef]
    [Google Scholar]
  6. Brzywczy J., Sieânko M., Kucharska A., Paszewski A. 2002; Sulphur amino acid synthesis in Schizosaccharomyces pombe represents a specific variant of sulphur metabolism in fungi. Yeast 19:29–35 [CrossRef]
    [Google Scholar]
  7. Bulmer G. S., Sans M. D., Gunn C. M. 1967; Cryptococcus neoformans. I. Nonencapsulated mutants. J Bacteriol 94:1475–1479
    [Google Scholar]
  8. Cox G. M., Mukherjee J., Cole G. T., Casadevall A., Perfect J. R. 2000; Urease as a virulence factor in experimental cryptococcosis. Infect Immun 68:443–448 [CrossRef]
    [Google Scholar]
  9. Cruz M. C., Del Poeta M., Wang P. 7 other authors 2000; Immunosuppressive and nonimmunosuppressive cyclosporine analogs are toxic to the opportunistic fungal pathogen Cryptococcus neoformans via cyclophilin-dependent inhibition of calcineurin. Antimicrob Agents Chemother 44:143–149 [CrossRef]
    [Google Scholar]
  10. Cruz M. C., Goldstein A. L., Blankenship J. R., Del Poeta M., Davis D., Cardenas M. E., Perfect J., McCusker J. H., Heitman J. 2002; Calcineurin is essential for survival during membrane stress in Candida albicans. EMBO J 21:546–559 [CrossRef]
    [Google Scholar]
  11. Davidson R. C., Blankenship J. R., Kraus P. R., de Jesus Berrios M., Hull C. M., D'Souza C., Wang P., Heitman J. 2002; A PCR-based strategy to generate integrative targeting alleles with large regions of homology. Microbiology 148:2607–2615
    [Google Scholar]
  12. Debono M., Gordee R. S. 1994; Antibiotics that inhibit fungal cell wall development. Annu Rev Microbiol 48:471–497 [CrossRef]
    [Google Scholar]
  13. Del Poeta M., Cruz M. C., Cardenas M. E., Perfect J. R., Heitman J. 2000; Synergistic antifungal activities of bafilomycin A1, fluconazole, and the pneumocandin MK-0991/caspofungin acetate (L-743,873) with calcineurin inhibitors FK506 and L-685,818 against Cryptococcus neoformans. Antimicrob Agents Chemother 44:739–746 [CrossRef]
    [Google Scholar]
  14. Fromtling R. A., Shadomy H. J., Jacobson E. S. 1982; Decreased virulence in stable, acapsular mutants of Cryptococcus neoformans. Mycopathologia 79:23–29 [CrossRef]
    [Google Scholar]
  15. Galgiani J. N., Lewis M. L. 1997; In vitro studies of activities of the antifungal triazoles SCH56592 and itraconazole against Candida albicans, Cryptococcus neoformans, and other pathogenic yeasts. Antimicrob Agents Chemother 41:180–183
    [Google Scholar]
  16. Gâomez B. L., Nosanchuk J. D. 2003; Melanin and fungi. Curr Opin Infect Dis 16:91–96 [CrossRef]
    [Google Scholar]
  17. Ghannoum M. A., Ibrahim A. S., Fu Y., Shafiq M. C., Edwards J. E. Jr, Criddle R. S. 1992; Susceptibility testing of Cryptococcus neoformans: a microdilution technique. J Clin Microbiol 30:2881–2886
    [Google Scholar]
  18. Gonzâalez J. C., Banerjee R. V., Huang S., Sumner J. S., Matthews R. G. 1992; Comparison of cobalamin-independent and cobalamin-dependent methionine synthases from Escherichia coli: two solutions to the same chemical problem. Biochemistry 31:6045–6056 [CrossRef]
    [Google Scholar]
  19. Granger D. L., Perfect J. R., Durack D. T. 1985; Virulence of Cryptococcus neoformans. Regulation of capsule synthesis by carbon dioxide. J Clin Invest 76:508–516 [CrossRef]
    [Google Scholar]
  20. Hatanaka H., Ariga N., Nagai J., Katsuki H. 1974; Accumulation of a sterol intermediate during reaction in the presence of homocysteine with cell-free extract of yeast. Biochem Biophys Res Commun 60:787–793 [CrossRef]
    [Google Scholar]
  21. Hoffman C. S., Winston F. 1987; A ten-minute DNA preparation from yeast efficiently releases autonomous plasmids for transformation of Escherichia coli. Gene 57:267–272 [CrossRef]
    [Google Scholar]
  22. Jakubowski H. 1991; Proofreading in vivo: editing of homocysteine by methionyl-tRNA synthetase in the yeast Saccharomyces cerevisiae. EMBO J 10:593–598
    [Google Scholar]
  23. Jakubowski H. 2002; The determination of homocysteine-thiolactone in biological samples. Anal Biochem 308:112–119 [CrossRef]
    [Google Scholar]
  24. Jakubowski H. 2004; Molecular basis of homocysteine toxicity in humans. Cell Mol Life Sci 61:470–487 [CrossRef]
    [Google Scholar]
  25. Kacprzak M. M., Lewandowska I., Matthews R. G., Paszewski A. 2003; Transcriptional regulation of methionine synthase by homocysteine and choline in Aspergillus nidulans. Biochem J 376:517–524 [CrossRef]
    [Google Scholar]
  26. Kingsbury J. M., Yang Z., Ganous T. M., Cox G. M., McCusker J. H. 2004a; Cryptococcus neoformans Ilv2p confers resistance to sulfometuron methyl and is required for survival at 37 °C and in vivo. Microbiology 150:1547–1558 [CrossRef]
    [Google Scholar]
  27. Kingsbury J. M., Yang Z., Ganous T. M., Cox G. M., McCusker J. H. 2004b; Novel chimeric spermidine synthase-saccharopine dehydrogenase gene (SPE3-LYS9) in the human pathogen Cryptococcus neoformans. Eukaryot Cell 3:752–763 [CrossRef]
    [Google Scholar]
  28. Kwon-Chung K. J., Bennett J. E. 1992; Cryptococcus. In Medical Mycology pp. 397–446 Edited by Cann C. Philadelphia, PA: Lea & Febiger;
    [Google Scholar]
  29. Kwon-Chung K. J., Rhodes J. C. 1986; Encapsulation and melanin formation as indicators of virulence in Cryptococcus neoformans. Infect Immun 51:218–223
    [Google Scholar]
  30. Maesaki S., Marichal P., Ashraf Hossain M., Sanglard D., Bossche H. V., Kohno S. 1998; Synergistic effects of tacrolimus and azole antifungal agents against azole-resistant Candida albicans strains. J Antimicrob Chemother 42:747–753 [CrossRef]
    [Google Scholar]
  31. Marchetti O., Moreillon P., Glauser M. P., Bille J., Sanglard D. 2000a; Potent synergism of the combination of fluconazole and cyclosporine in Candida albicans. Antimicrob Agents Chemother 44:2373–2381 [CrossRef]
    [Google Scholar]
  32. Marchetti O., Entenza J. M., Sanglard D., Bille J., Glauser M. P., Moreillon P. 2000b; Fluconazole plus cyclosporine: a fungicidal combination effective against experimental endocarditis due to Candida albicans. Antimicrob Agents Chemother 44:2932–2938 [CrossRef]
    [Google Scholar]
  33. Marchetti O., Moreillon P., Entenza J. M., Vouillamoz J., Glauser M. P., Bille J., Sanglard D. 2003; Fungicidal synergism of fluconazole and cyclosporine in Candida albicansis not dependent on multidrug efflux transporters encoded by the CDR1, CDR2, CaMDR1, and FLU1 genes. Antimicrob Agents Chemother 47:1565–1570 [CrossRef]
    [Google Scholar]
  34. Maresca B., Kobayashi G. S. 1989; Dimorphism in Histoplasma capsulatum: a model for the study of cell differentiation in pathogenic fungi. Microbiol Rev 53:186–209
    [Google Scholar]
  35. Marzluf G. A. 1997; Molecular genetics of sulfur assimilation in filamentous fungi and yeast. Annu Rev Microbiol 51:73–96 [CrossRef]
    [Google Scholar]
  36. McCammon M. T., Parks L. W. 1981; Inhibition of sterol transmethylation by S-adenosylhomocysteine analogs. J Bacteriol 145:106–112
    [Google Scholar]
  37. McDade H. C., Cox G. M. 2001; A new dominant selectable marker for use in Cryptococcus neoformans. Med Mycol 39:151–154 [CrossRef]
    [Google Scholar]
  38. Medoff G., Painter A., Kobayashi G. S. 1987; Mycelial- to yeast-phase transitions of the dimorphic fungi Blastomyces dermatitidis and Paracoccidioides brasiliensis. J Bacteriol 169:4055–4060
    [Google Scholar]
  39. Odom A., Del Poeta M., Perfect J., Heitman J. 1997a; The immunosuppressant FK506 and its nonimmunosuppressive analog L-685,818 are toxic to Cryptococcus neoformans by inhibition of a common target protein. Antimicrob Agents Chemother 41:156–161
    [Google Scholar]
  40. Odom A., Muir S., Lim E., Toffaletti D. L., Perfect J., Heitman J. 1997b; Calcineurin is required for virulence of Cryptococcus neoformans. EMBO J 16:2576–2589 [CrossRef]
    [Google Scholar]
  41. Onyewu C., Blankenship J. R., Del Poeta M., Heitman J. 2003; Ergosterol biosynthesis inhibitors become fungicidal when combined with calcineurin inhibitors against Candida albicans, Candida glabrata, and Candida krusei. Antimicrob Agents Chemother 47:956–964 [CrossRef]
    [Google Scholar]
  42. Parks L. W., Casey W. M. 1995; Physiological implications of sterol biosynthesis in yeast. Annu Rev Microbiol 49:95–116 [CrossRef]
    [Google Scholar]
  43. Perfect J. R., Toffaletti D. L., Rude T. H. 1993; The gene encoding phosphoribosylaminoimidazole carboxylase (ADE2) is essential for growth of Cryptococcus neoformans in cerebrospinal fluid. Infect Immun 61:4446–4451
    [Google Scholar]
  44. Pieniazek N., Stepieân P. P., Paszewski A. 1973; An Aspergillus nidulans mutant lacking cystathionine synthase: identity of l-serine sulfhydrylase with cystathionine synthase and its distinctness from o-acetyl-l-serine sulfhydrylase. Biochim Biophys Acta 297:37–47 [CrossRef]
    [Google Scholar]
  45. Rose M. D., Winston F., Hieter P. 1990 Methods in Yeast Genetics: a Laboratory Course Manual Cold Spring Harbor, NY: Cold Spring Harbor Laboratory;
    [Google Scholar]
  46. Sambrook J., Fritsch E. F., Maniatis T. 1989 Molecular Cloning; a Laboratory Manual, 2nd edn. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory;
    [Google Scholar]
  47. Sanglard D. 2002; Resistance of human fungal pathogens to antifungal drugs. Curr Opin Microbiol 5:379–385 [CrossRef]
    [Google Scholar]
  48. Sanglard D., Ischer F., Marchetti O., Entenza J. M., Bille J. 2003; Calcineurin A of Candida albicans: involvement in antifungal tolerance, cell morphogenesis and virulence. Mol Microbiol 48:959–976 [CrossRef]
    [Google Scholar]
  49. Sheehan D. J., Hitchcock C. A., Sibley C. M. 1999; Current and emerging azole antifungal agents. Clin Microbiol Rev 12:40–79
    [Google Scholar]
  50. Thomas D., Surdin-Kerjan Y. 1997; Metabolism of sulfur amino acids in Saccharomyces cerevisiae. Microbiol Mol Biol Rev 61:503–532
    [Google Scholar]
  51. Toffaletti D. L., Rude T. H., Johnston S. A., Durack D. T., Perfect J. R. 1993; Gene transfer in Cryptococcus neoformans by use of biolistic delivery of DNA. J Bacteriol 175:1405–1411
    [Google Scholar]
  52. Vallim M. A., Fernandes L., Alspaugh J. 2004; The RAM1 gene encoding a protein-farnesyltransferaseβ-subunit homolog is essential in Cryptococcus neoformans. Microbiology 150:1925–1935 [CrossRef]
    [Google Scholar]
  53. White T. C., Marr K. A., Bowden R. A. 1998; Clinical, cellular, and molecular factors that contribute to antifungal drug resistance. Clin Microbiol Rev 11:382–402
    [Google Scholar]
  54. Yang Z., Pascon R. C., Alspaugh A., Cox G. M., McCusker J. H. 2002; Molecular and genetic analysis of the Cryptococcus neoformans MET3 gene and a met3 mutant. Microbiology 148:2617–2625
    [Google Scholar]
  55. Zaragoza O., Fries B. C., Casadevall A. 2003; Induction of capsule growth in Cryptococcus neoformans by mammalian serum and CO2. Infect Immun 71:6155–6164 [CrossRef]
    [Google Scholar]
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