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Volume 147, Issue 7

Trehalose is required for the acquisition of tolerance to a variety of stresses in the filamentous fungus

The GenBank accession number for the sequence reported in this paper is AF043230.

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Abstract

Trehalose is a non-reducing disaccharide found at high concentrations in conidia and rapidly degraded upon induction of conidial germination. Furthermore, trehalose is accumulated in response to a heat shock or to an oxidative shock. The authors have characterized the gene encoding trehalose-6-phosphate synthase, which catalyses the first step in trehalose biosynthesis. Expression of in a mutant revealed that the gene product is a functional equivalent of the yeast Tps1 trehalose-6-phosphate synthase. The null mutant does not produce trehalose during conidiation or in response to various stress conditions. While germlings of the mutant show an increased sensitivity to moderate stress conditions (growth at 45 °C or in the presence of 2 mM HO), they display a response to severe stress (60 min at 50 °C or in the presence of 100 mM HO) similar to that of wild-type germlings. Furthermore, conidia of the mutant show a rapid loss of viability upon storage. These results are consistent with a role of trehalose in the acquisition of stress tolerance. Inactivation of the gene also results in increased steady-state levels of sugar phosphates but does not prevent growth on rapidly metabolizable carbon sources (glucose, fructose) as seen in . This suggests that trehalose 6-phosphate is a physiological inhibitor of hexokinase but that this control is not essential for proper glycolytic flux in . Interestingly, transcription is not induced in response to heat shock or during conidiation, indicating that trehalose accumulation is probably due to a post-translational activation process of the trehalose 6-phosphate synthase.

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  • Accepted:
  • Revised:
  • Published Online:
Keyword(s): 5-FOA, 5-fluoro-orotic acid , EST, expressed sequence tag , glycolysis , heat stress , hexokinase , oxidative stress , spore germination , T6P, trehalose 6-phosphate , T6PP, trehalose-6-phosphate phosphatase , T6PS, trehalose-6-phosphate synthase and trehalose 6-phosphate synthase
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2001-07-01
2024-04-24
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References

  1. Arguelles J. C. 2000; Physiological roles of trehalose in bacteria and yeasts: a comparative analysis. Arch Microbiol 174:217–224 [CrossRef]
     [Google Scholar]
  2. Arisan-Atac I., Wolschek M. F., Kubicek C. P. 1996; Trehalose-6-phosphate synthase A affects citrate accumulation by Aspergillus niger under conditions of high glycolytic flux. FEMS Microbiol Lett 140:77–83 [CrossRef]
     [Google Scholar]
  3. Ausubel F. M., Brent R., Kingston R. E., Moore D. D., Seidman J. G., Smith J. A., Struhl K. 1992 Short Protocols in Molecular Biology. New York: Wiley;
     [Google Scholar]
  4. Bell W., Klaassen P., Ohnacker M., Boller T., Herweijer M., Schoppink P., Wiemken A, Van der Zee P. 1992; Characterization of the 56-kDa subunit of yeast trehalose-6-phosphate synthase and cloning of its gene reveal its identity with the product of CIF1, a regulator of carbon catabolite inactivation. Eur J Biochem 209:951–959 [CrossRef]
     [Google Scholar]
  5. Bell W., Sun W., Hohmann S., Wera S., Reinders A., Wiemken A., Thevelein J. M, de Virgilio C. 1998; Composition and functional analysis of the Saccharomyces cerevisiae trehalose synthase complex. J Biol Chem 273:33311–33319 [CrossRef]
     [Google Scholar]
  6. Blazquez M. A., Lagunas R., Gancedo C., Gancedo J. M. 1993; Trehalose-6-phosphate, a new regulator of yeast glycolysis that inhibits hexokinases. FEBS Lett 329:51–54 [CrossRef]
     [Google Scholar]
  7. Blazquez M. A., Stucka R., Feldmann H., Gancedo C. 1994; Trehalose-6-P synthase is dispensable for growth on glucose but not for spore germination in Schizosaccharomyces pombe. J Bacteriol 1763895–3902
     [Google Scholar]
  8. Bonini B. M., Van Vaeck C., Larsson C., Gustafsson L., Ma P., Winderickx J., Van Dijck P., Thevelein J. M. 2000; Expression of Escherichia coli otsA in a Saccharomyces cerevisiae tps1 mutant restores trehalose 6-phosphate levels and partly restores growth and fermentation with glucose and control of glucose influx into glycolysis. Biochem J 350:261–268 [CrossRef]
     [Google Scholar]
  9. Borgia P. T., Miao Y., Dodge C. L. 1996; The orlA gene from Aspergillus nidulans encodes a trehalose-6-phosphate phosphatase necessary for normal growth and chitin synthesis at elevated temperatures. Mol Microbiol 20:1287–1296 [CrossRef]
     [Google Scholar]
  10. Brody H., Griffith J., Cuticchia A. J., Arnold J., Timberlake W. E. 1991; Chromosome-specific recombinant DNA library from the fungus Aspergillus nidulans. Nucleic Acids Res 19:3105–3109 [CrossRef]
     [Google Scholar]
  11. Chen D. C., Yang B. C., Kuo T. T. 1992; One-step transformation of yeast in stationary phase. Curr Genet 21:83–84 [CrossRef]
     [Google Scholar]
  12. Devereux J., Haeberli P., Smithies O. 1984; A comprehensive set of sequence analysis programs for the VAX. Nucleic Acids Res 12:387–395 [CrossRef]
     [Google Scholar]
  13. Elbein A. D. 1974; The metabolism of α,α-trehalose. Adv Carbohydr Chem Biochem 30:256–277
     [Google Scholar]
  14. d’Enfert C. 1996; Selection of multiple disruption events in Aspergillus fumigatus using the orotidine-5′-decarboxylase gene, pyrG as a unique transformation marker. Curr Genet 30:76–82 [CrossRef]
     [Google Scholar]
  15. d’Enfert C., Fontaine T. 1997; Molecular characterization of the Aspergillus nidulans treA gene encoding an acid trehalase required for growth on trehalose. Mol Microbiol 24:203–216 [CrossRef]
     [Google Scholar]
  16. d’Enfert C., Bonini B. M., Zapella P. D. A., Fontaine T, da Silva A. M., Terenzi H. F. 1999; Neutral trehalases catalyse intracellular trehalose breakdown in the filamentous fungi Aspergillus nidulans and Neurospora crassa. Mol Microbiol 32:471–484 [CrossRef]
     [Google Scholar]
  17. Girardin H., Morrow B., Soll D, Latgé J.-P., Skirantha T. 1993; Development of DNA probes for fingerprinting Aspergillus fumigatus. J Clin Microbiol 31:1547–1554
     [Google Scholar]
  18. Hanahan D., Jessee J., Bloom F. R. 1991; Plasmid transformation of Escherichia coli and other bacteria. Methods Enzymol 204:63–113
     [Google Scholar]
  19. Hohmann S., Bell W., Neves M. J., Valckx D., Thevelein J. M. 1996; Evidence for trehalose-6-phosphate-dependent and -independent mechanisms in the control of sugar influx into yeast glycolysis. Mol Microbiol 20:981–991 [CrossRef]
     [Google Scholar]
  20. Hounsa C.-G., Vincent E., Thevelein J. M., Hohmann S., Prior B. 1998; Role of trehalose in survival of Saccharomyces cerevisiae under osmotic stress. Microbiology 144:671–680 [CrossRef]
     [Google Scholar]
  21. Kopp M., Holzer H, Müller H. 1993; Molecular analysis of the neutral trehalase gene from Saccharomyces cerevisiae. J Biol Chem 268:4766–4774
     [Google Scholar]
  22. Lewis J. G., Learmonth R. P., Watson K. 1995; Induction of heat, freezing and salt tolerance by heat and salt shock in Saccharomyces cerevisiae. Microbiology 141687–694 [CrossRef]
     [Google Scholar]
  23. Lingappa B. T., Sussman A. S. 1959; Endogenous substrates of dormant, activated and germinating ascospores of Neurospora tetrasperma. Plant Physiol 34:466–473 [CrossRef]
     [Google Scholar]
  24. Llorente B., Fairhead C., Dujon B. 1999; Genetic redundancy and gene fusion in the genome of the baker’s yeast Saccharomyces cerevisiae : functional characterization of a three-member gene family involved in the thiamine biosynthetic pathway. Mol Microbiol 32:1140–1152 [CrossRef]
     [Google Scholar]
  25. Luyten K, de Koning W., Tesseur I., Ruiz M. C., Ramos J., Cobbaert P., Thevelein J. M., Hohmann S. 1993; Disruption of the Kluyveromyces lactis GGS1 gene causes inability to grow on glucose and fructose and is suppressed by mutations that reduce sugar uptake. Eur J Biochem 217:701–713 [CrossRef]
     [Google Scholar]
  26. Mol P. C., Mouyna I., d’Enfert C. 1998; A rapid procedure for the extraction of genomic DNA from intact Aspergillus spores. Fungal Genet Newsl 45:28–29
     [Google Scholar]
  27. Neves M. J., Terenzi H. F., Leone F. A., Jorge J. A. 1994; Quantification of trehalose in biological samples with a conidial trehalase from the thermophilic fungus Humicola grisea var.thermoidea. World J Microbiol Biotechnol 10:17–19 [CrossRef]
     [Google Scholar]
  28. Neves M. J., Hohmann S., Bell W. 7 other authors 1995; Control of glucose influx into glycolysis and pleiotropic effects studied in different isogenic sets of Saccharomyces cerevisiae mutants in trehalose biosynthesis. Curr Genet 27:110–122 [CrossRef]
     [Google Scholar]
  29. Noventa-Jordao M. A., Couto R. M., Goldman M. H., Aguirre J., Iyer S., Caplan A., Terenzi H. F., Goldman G. H. 1999; Catalase activity is necessary for heat-shock recovery in Aspergillus nidulans germlings. Microbiology 145:3229–3234
     [Google Scholar]
  30. Nwaka S., Mechler B., Destruelle M., Holzer H. 1995; Phenotypic features of trehalase mutants in Saccharomyces cerevisiae. FEBS Lett 360:286–290 [CrossRef]
     [Google Scholar]
  31. Osmani S. A., May G. S., Morris R. N. 1987; Regulation of the mRNA levels of nimA , a gene required for the G2-M transition in Aspergillus nidulans. J Cell Biol 104:1495–1504 [CrossRef]
     [Google Scholar]
  32. Panneman H., Ruijter G. J. G., Visser J, van den Broeck H. C. 1998; Cloning and biochemical characterization of Aspergillus niger hexokinase. The enzyme is strongly inhibited by physiological concentrations of trehalose-6-phosphate. Eur J Biochem 258:223–232 [CrossRef]
     [Google Scholar]
  33. Prade R. A., Griffith J., Kochut K., Arnold J., Timberlake W. E. 1997; In vitro reconstruction of the Aspergillus (=Emericella) nidulans genome. Proc Natl Acad Sci USA 94:14564–14569 [CrossRef]
     [Google Scholar]
  34. Reinders A., Burckert N., Hohmann S., Thevelein J. M., Boller T., Weimken A., De Virgilio C. 1997; Structural analysis of the subunits of the trehalose-6-phosphate synthase/phosphatase complex in Saccharomyces cerevisiae and their function during heat shock. Mol Microbiol 24:687–695 [CrossRef]
     [Google Scholar]
  35. Reinders A., Burckert N., Boller T., Wiemken A., De Virgilio C. 1998; Saccharomyces cerevisiae cAMP-dependent protein kinase controls entry into stationary phase through the Rim15p protein kinase. Genes Dev 12:2943–2955 [CrossRef]
     [Google Scholar]
  36. Ribeiro M. J. S., Reinders A., Boller T., Wiemken A, de Virgilio C. 1998; Trehalose synthesis is important for the acquisition of thermotolerance in Schizosaccharomyces pombe. Mol Microbiol 25. 571–581
  37. Ruijter G. J. G., Visser J. 1996; Determination of intermediary metabolites in Aspergillus niger. J Microbiol Methods 25:295–302 [CrossRef]
     [Google Scholar]
  38. Ruijter G. J. G., Panneman H, van den Broeck H. C., Bennett J. M., Visser J. 1996; Characterisation of the Aspergillus nidulans fr A1 mutant: hexose phosphorylation and apparent lack of involvement of hexokinase in glucose repression. FEMS Microbiol Lett 139:223–228 [CrossRef]
     [Google Scholar]
  39. Sambrook J., Fritsch E. F., Maniatis T. 1989 Molecular Cloning: a Laboratory Manual Cold Spring Harbor, NY: Cold Spring Harbor Laboratory;
     [Google Scholar]
  40. Sanger F., Nicklen S., Coulson R. A. 1977; DNA sequencing with chain terminating inhibitors. Proc Natl Acad Sci USA 74:5463–5467 [CrossRef]
     [Google Scholar]
  41. Sherman F. 1991; Getting started with yeast. Methods Enzymol 194:3–21
     [Google Scholar]
  42. Thevelein J. M. 1984; Regulation of trehalose mobilization in fungi. Microbiol Rev 48:42–59
     [Google Scholar]
  43. Thevelein J. M. 1996; Regulation of trehalose metabolism and its relevance to cell growth and function. . In The Mycota III. Biochemistry and Molecular Biology pp 395–420 Edited by Brambl R., Marzluf G. A. Berlin: Springer;
     [Google Scholar]
  44. Thevelein J. M., Hohmann S. 1995; Trehalose synthase: guard to the gate of glycolysis in yeast. Trends Biochem Sci 20:3–9 [CrossRef]
     [Google Scholar]
  45. Thevelein J. M., de Winde J. H. 1999; Novel sensing mechanisms and targets for the cAMP-protein kinase A pathway in the yeast Saccharomyces cerevisiae. Mol Microbiol 33:904–918 [CrossRef]
     [Google Scholar]
  46. Thomas B. J., Rothstein R. 1989; Elevated recombination rates in transcriptionally active DNA. Cell 56:619–630 [CrossRef]
     [Google Scholar]
  47. Van Aelst L., Hohmann S., Bulaya B. 7 other authors 1993; Molecular cloning of a gene involved in glucose sensing in the yeast Saccharomyces cerevisiae. Mol Microbiol 8:927–943 [CrossRef]
     [Google Scholar]
  48. Van Laere A. 1989; Trehalose, reserve and/or stress metabolite?. FEMS Microbiol Rev 63:201–210
     [Google Scholar]
  49. de Virgilio C., Bückert N., Bell W., Jenö P., Boller T., Wiemken A. 1993; Disruption of TPS2 , the gene encoding the 100 kDa subunit of the trehalose-6-phosphate synthase/phosphatase complex in Saccharomyces cerevisiae , causes accumulation of trehalose-6-phosphate and loss of trehalose-6-phosphate phosphatase activity. Eur J Biochem 212:315–323 [CrossRef]
     [Google Scholar]
  50. Vuorio O. E., Kalkkinen N., Londesborough J. 1993; Cloning of two related genes encoding the 56-kDa and 123-kDa subunits of trehalose synthase from the yeast Saccharomyces cerevisiae. Eur J Biochem 216:849–861 [CrossRef]
     [Google Scholar]
  51. Weidner G., d’Enfert C., Koch A., Mol P. C., Brakhage A. A. 1998; Development of a homologous transformation system for the pathogenic fungus Aspergillus fumigatus based on the pyrG gene encoding orotidine monophosphate decarboxylase. Curr Genet 33:378–385 [CrossRef]
     [Google Scholar]
  52. Wiemken A. 1990; Trehalose in yeast, stress protectant rather than reserve carbohydrate. Antonie Leeuwenhoek 58:209–217 [CrossRef]
     [Google Scholar]
  53. Winderickx J, de Winde J. H., Crauwels M., Hino A., Hohmann S., Van Dijck P., Thevelein J. M. 1996; Regulation of genes encoding subunits of the trehalose synthase complex in Saccharomyces cerevisiae : novel variations of STRE-mediated transcription control?. Mol Gen Genet 252:470–482
     [Google Scholar]
  54. Wolschek M. F., Kubicek C. P. 1997; The filamentous fungus Aspergillus niger contains two ‘‘differentially regulated’’ trehalose-6-phosphate synthase-encoding genes,tpsA and tpsB. . J Biol Chem 272:2729–2735 [CrossRef]
     [Google Scholar]
  55. Woodcock D. M., Crowther P. J., Doherty J., Jefferson S., DeCruz E., Noyer-Weidner M., Smith S. S., Michael M. Z., Graham M. W. 1989; Quantitative evaluation of Escherichia coli host strains for tolerance to cytosine methylation in plasmid and phage recombinants. Nucleic Acids Res 17:3469–3478 [CrossRef]
     [Google Scholar]
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Trehalose is required for the acquisition of tolerance to a variety of stresses in the filamentous fungus Aspergillus nidulansThe GenBank accession number for the sequence reported in this paper is AF043230.
Microbiology 147, 1851 (2001); https://doi.org/10.1099/00221287-147-7-1851
/content/journal/micro/10.1099/00221287-147-7-1851
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