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Journal of Microbiology

, Volume 57, Issue 7, pp 606–617 | Cite as

Alcohol dehydrogenase 1 participates in the Crabtree effect and connects fermentative and oxidative metabolism in the Zygomycete Mucor circinelloides

  • Rosa Angélica Rangel-Porras
  • Sharel P. Díaz-Pérez
  • Juan Manuel Mendoza-Hernández
  • Pamela Romo-Rodríguez
  • Viridiana Alejandre-Castañeda
  • Marco I. Valle-Maldonado
  • Juan Carlos Torres-Guzmán
  • Gloria Angélica González-Hernández
  • Jesús Campos-Garcia
  • José Arnau
  • Víctor Meza-Carmen
  • J. Félix Gutiérrez-CoronaEmail author
Microbial Physiology and Biochemistry
  • 46 Downloads

Abstract

Mucor circinelloides is a dimorphic Zygomycete fungus that produces ethanol under aerobic conditions in the presence of glucose, which indicates that it is a Crabtree-positive fungus. To determine the physiological role of the alcohol dehydrogenase (ADH) activity elicited under these conditions, we obtained and characterized an allyl alcohol-resistant mutant that was defective in ADH activity, and examined the effect of adh mutation on physiological parameters related to carbon and energy metabolism. Compared to the Adh+ strain R7B, the ADH-defective (Adh-) strain M5 was unable to grow under anaerobic conditions, exhibited a considerable reduction in ethanol production in aerobic cultures when incubated with glucose, had markedly reduced growth capacity in the presence of oxygen when ethanol was the sole carbon source, and exhibited very low levels of NAD+-dependent alcohol de-hydrogenase activity in the cytosolic fraction. Further characterization of the M5 strain showed that it contains a 10-bp deletion that interrupts the coding region of the adhl gene. Complementation with the wild-type allele adh1+ by transformation of M5 remedied all the defects caused by the adh1 mutation. These findings indicate that in M. circinelloides, the product of the adh1 gene mediates the Crabtree effect, and can act as either a fermentative or an oxidative enzyme, depending on the nutritional conditions, thereby participating in the association between fermentative and oxidative metabolism. It was found that the spores of M. circinelloides possess low mRNA levels of the ethanol assimilation genes (adl2 and acs2), which could explain their inability to grow in the alcohol.

Keywords

Mucor circinelloides ADH1 enzyme Crabtree effect fermentative and oxidative metabolism 

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Notes

Acknowledgements

This work was supported by grants 29078N, 41590, and 167071 from SEP-CONACyT, México. RARP, SPDP, VAC, MIVM, and JMMH each received a fellowship from CONACyT, México. We thank María E. Cardenas for critical reading of the manuscript.

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References

  1. Acevedo-Aguilar, F.J., Espino-Saldaña, A.E., León-Rodríguez, I.L., Ávila-Rodríguez, M., Wrobel, K., Wrobel, K., Lappe, P., Ulloa, M., and Gutiérrez-Corona, J.F. 2006. Hexavalent chromium removal in vitro and from industrial wastes, using chromate-resistant strains of filamentous fungi indigenous to contaminated wastes. Can. J. Microbiol. 52, 809–815.CrossRefGoogle Scholar
  2. Bartnicki-García S. 1963. Symposium on biochemical bases of morphogenesis in fungi III Mold-yeast dimorphism of Mucor. Bacteriol. Rev. 27, 293–304.Google Scholar
  3. Bartnicki-Garcia, S., Nelson, N., and Cota-Robles, E. 1968. Electron microscopy of spore germination and cell wall formation in Mucor rouxii. Arch. Mikrobiol. 63, 242–255.CrossRefGoogle Scholar
  4. Bartnicki-García, S. and Nickerson, W.J. 1962. Nutrition, growth, and morphogenesis of Mucor rouxii. J. Bacteriol. 84, 841–858.Google Scholar
  5. Bergmeyer, H.U. 1983. Reagents for enzymatic analysis, pp. 139. In Bergmeyer, H.U. (ed.), Methods of enzymatic analysis. Verlag Chemie, Weinheim, Germany.Google Scholar
  6. Chayakulkeeree, M., Ghannoum, M., and Perfect, J. 2006. Zygo-mycosis: the re-emerging fungal infection. Eur. J. Clin. Microbiol. Infect. Dis. 25, 215–229.CrossRefGoogle Scholar
  7. Corrales-Escobosa, A.R., Rangel-Porras, R.A., Meza-Carmen, V., Gonzalez-Hernandez, G.A., Torres-Guzman, J.C., Wrobel, K., Wrobel, K., Roncero, M.I.G., and Gutierrez-Corona, J.F. 2011. Fusarium oxysporum Adh1 has dual fermentative and oxidative functions and is involved in fungal virulence in tomato plants. Fungal Genet. Biol. 48, 886–895.CrossRefGoogle Scholar
  8. de Kok, S., Kozak, B.U., Pronk, J.T., and van Maris, A.J.A. 2012. Energy coupling in Saccharomyces cerevisiae: selected opportunities for metabolic engineering. FEMS Yeast Res. 12, 387–397.CrossRefGoogle Scholar
  9. de Smidt, O., du Preez, J.C., and Albertyn, J. 2008. The alcohol dehydrogenases of Saccharomyces cerevisiae: a comprehensive review. FEMS Yeast Res. 8, 967–978.CrossRefGoogle Scholar
  10. Ferreira, J.A., Lennartsson, P.R., Edebo, L., and Taherzadeh, M.J. 2013. Zygomycetes-based biorefinery: Present status and future prospects. Bioresour. Technol. 135, 523–532.CrossRefGoogle Scholar
  11. Freeling, M. and Bennett, D.C. 1985. Maize Adh1. Annu. Rev. Genet. 19, 297–323.CrossRefGoogle Scholar
  12. Hagman, A., Säll, T., Compagno, C., and Piskur, J. 2013. Yeast “Make-Accumulate-Consume” life strategy evolved as a multi-step process that predates the whole genome duplication. PLoS One 8, e68734.CrossRefGoogle Scholar
  13. Ibrahim, A.S. and Spellberg, B. 2006. Zygomycetes as agents of infectious disease in humans, pp. 429–440. In Heitman, J., Filler, S.G., Edwards, Jr. J.E., and Mitchell, A.P. (eds.), Molecular principles of fungal pathogenesis. American Society for Microbiology Press, Washington, D.C., USA.Google Scholar
  14. Jacobs, M., Dolferus, R., and Van den Bossche, D. 1988. Isolation and biochemical analysis of ethyl methanesulfonate induced alcohol dehydrogenase null mutants of Arabidopsis thaliana (L.) Heynh. Biochem. Genet. 26, 105–122.CrossRefGoogle Scholar
  15. Karimi, K. and Zamani, A. 2013. Mucor indicus: biology and industrial application perspectives: a review. Biotechnol. Adv. 31, 466–481.CrossRefGoogle Scholar
  16. Khan, Z.U., Ahmad, S., Brazda, A., and Chandy, R. 2009. Mucor circinelloides as a cause of invasive maxillofacial zygomycosis: an emerging dimorphic pathogen with reduced susceptibility to posaconazole. J. Clin. Microbiol. 47, 1244–1248.CrossRefGoogle Scholar
  17. Lee, K.L., Buckley, H.R., and Campbell, C.C. 1975. An aminoacid liquid synthetic medium for the development of mycelial and yeast forms of Candida albicans. Sabouraudia 13, 148–153.CrossRefGoogle Scholar
  18. Lee, S.C., Li, A., Calo, S., and Heitman, J. 2013. Calcineurin plays key roles in the dimorphic transition and virulence of the human pathogenic zygomycete Mucor circinelloides. PLoS Pathog. 9, e1003625.CrossRefGoogle Scholar
  19. Linz, J.E. and Orlowski, M. 1982. Stored mRNA in sporangiospores of the fungus Mucor recemosus. J. Bacteriol. 150, 1138–1144.Google Scholar
  20. Lopez-Alvarez, A., Díaz-Pérez, A.L., Sosa-Aguirre, C., Macías-Rod-riguez, L., and Campos-García, J. 2012. Ethanol yield and volatile compound content in fermentation of agave must by Kluyvero-myces marxianus UMPe-1 comparing with Saccharomyces cerevisiae baker’s yeast used in tequila production. J. Biosci. Bioeng. 113, 614–618.CrossRefGoogle Scholar
  21. Lübbehüsen, T.L., Nielsen, J., and Mclntyre, M. 2004. Aerobic and anaerobic ethanol production by Mucor circinelloides during submerged growth. Appl. Microbiol. Biotechnol. 63, 543–548.CrossRefGoogle Scholar
  22. Lutstorf, U. and Megnet, R. 1968. Multiple forms of alcohol dehy-drogenase in Saccharomyces cerevisiae. I. Physiological control of ADH-2 and properties of ADH-2 and ADH-4. Arch. Biochem. Biophys. 126, 933–944.CrossRefGoogle Scholar
  23. McIntyre, M., Breum, J., Arnau, J., and Nielsen, J. 2002. Growth physiology and dimorphism of Mucor circinelloides (syn. racemosus) during submerged batch cultivation. Appl. Microbiol. Biotechnol. 58, 495–502.CrossRefGoogle Scholar
  24. Mendoza, L., Vilela, R., Voelz, K., Ibrahim, A.S., Voigt, K., and Lee, S.C. 2015. Human fungal pathogens of mucorales and entomo-phthorales. Cold Spring Harb. Perspect. Med. 5, a019562.CrossRefGoogle Scholar
  25. Nikolova, P. and Ward, O.P. 1991. Production of L-phenylacetyl carbinol by biotransformation: product and by-product formation and activities of the key enzymes in wild-type and ADH isoenzyme mutants of Saccharomyces cerevisiae. Biotechnol. Bioeng. 20, 493–498.CrossRefGoogle Scholar
  26. Orlowski, M. 1991. Mucor dimorphism. Microbiol. Rev. 55, 234–258.Google Scholar
  27. Orlowski, M. and Sypherd, P.S. 1978. Regultion of macromolecular synthesis during hyphal germ tube emergence from Mucor race-mosus sporangiospores. J. Bacteriol. 134, 76–83.Google Scholar
  28. Panagiotou, G., Villas-Boas, S.G., Christakopoulos, P., Nielsen, J., and Olsson, L. 2005. Intracellular metabolite profiling of Fusarium oxysporum converting glucose to ethanol. J. Biotechnol. 115, 425–434.CrossRefGoogle Scholar
  29. Pfeiffer, T. and Morley, A. 2014. An evolutionary perspective on the Crabtree effect. Front. Mol. Biosci. 1, 17.CrossRefGoogle Scholar
  30. Pronk. J.T., Yde Steensma, H., and Van Dijken, J.P. 1996. Pyruvate metabolism in Saccharomyces cerevisiae. Yeast 12, 1607–1633.CrossRefGoogle Scholar
  31. Rangel-Porras, R.A., Meza-Carmen, V., Martínez-Cadena, G., Torres-Guzmán, J.C. González-Hernandez, G.A., Arnau, J., and Gutiérrez-Corona, J.F. 2005. Molecular analysis of an NAD-dependent alcohol dehydrogenase from the zygomycete Mucor circinelloides. Mol. Genet. Genomics 274, 354–363.CrossRefGoogle Scholar
  32. Reid, M.F. and Fewson, C.A. 1994. Molecular characterization of microbial alcohol dehydrogenases. Crit. Rev. Microbiol. 20, 13–56.CrossRefGoogle Scholar
  33. Roncero, M.I.G. 1984. Enrichment method for the isolation of auxotrophic mutants of Mucor using the polyene antibiotic N-glycosyl-polifungin. Carlsberg Res. Commun. 49, 685–690.CrossRefGoogle Scholar
  34. Salcedo-Hernández, R. and Ruiz-Herrera, J. 1993. Isolation and characterization of a mycelial cytochrome aa3-deficient mutant and the role of mitochondria in dimorphism of Mucor rouxii. Exp. Mycol. 17, 142–154.CrossRefGoogle Scholar
  35. Sambrook, J., Fritsch, E.F., and Maniatis, T. 1989. Molecular cloning: a laboratory manual (2nd ed.). Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, USA.Google Scholar
  36. Sypherd, P., Borgia, P.T., and Paznokas, J.L. 1978. Biochemistry of dimorphism in the fungus Mucor. Adv. Microb. Physiol. 18, 67–104.CrossRefGoogle Scholar
  37. Torres-Guzmán, J.C., Arreola-García, G.A., Zazueta-Sandoval, R., Carrillo-Rayas, T., Martínez-Cadena, G., and Gutiérrez-Corona, F. 1994. Genetic evidence for independence between fermentative metabolism (ethanol accumulation) and yeast-cell development in the dimorphic fungus Mucor rouxii. Curr. Genet. 26, 166–171.CrossRefGoogle Scholar
  38. Valle-Maldonado, M.I., Jácome-Galarza, I.E., Gutiérrez-Corona, F., Ramírez-Díaz, M.I., Campos-García, J., and Meza-Carmen, V. 2015. Selection of reference genes for quantitative real time RT-PCR during dimorphism in the zygomycete Mucor circinel-loides. Mol. Biol. Rep. 42, 705–711.CrossRefGoogle Scholar
  39. Williamson, V.M., Long, M., and Theodoris, G. 1991. Isolation of Caenorhabditis elegans mutants lacking alcohol dehydrogenase activity. Biochem. Genet. 29, 313–323.CrossRefGoogle Scholar
  40. Wills, C. 1990. Regulation of sugar and ethanol metabolism in Saccharomyces cerevisiae. Crit. Rev. Biochem. Mol. Biol. 25, 245–280.CrossRefGoogle Scholar
  41. Wills, C. and Phelps, J. 1975. A technique for the isolation of yeast alcohol dehydrogenase mutants with altered substrate specificity. Arch. Biochem. Biophys. 167, 627–637.CrossRefGoogle Scholar
  42. Wolff, A.M. and Arnau, J. 2002. Cloning of glyceraldehyde-3-phos-phate dehydrogenase-encoding genes in Mucor circinelloides (Syn. racemosus) and use of the gpd1 promoter for recombinant protein production. Fungal Genet. Biol. 35, 21–29.CrossRefGoogle Scholar
  43. Zheng, Z., Luo, S., Li, X., Wu, X., Pan, L., and Jiang, S. 2009. Screening of allyl alcohol resistant mutant of Rhizopus oryzae and its fermentation characterization. Afr. J. Biotechnol. 8, 280–284.Google Scholar

Copyright information

© The Microbiological Society of Korea 2019

Authors and Affiliations

  • Rosa Angélica Rangel-Porras
    • 1
  • Sharel P. Díaz-Pérez
    • 2
  • Juan Manuel Mendoza-Hernández
    • 1
  • Pamela Romo-Rodríguez
    • 1
  • Viridiana Alejandre-Castañeda
    • 2
  • Marco I. Valle-Maldonado
    • 2
    • 3
  • Juan Carlos Torres-Guzmán
    • 1
  • Gloria Angélica González-Hernández
    • 1
  • Jesús Campos-Garcia
    • 2
  • José Arnau
    • 4
  • Víctor Meza-Carmen
    • 2
  • J. Félix Gutiérrez-Corona
    • 1
    Email author
  1. 1.Department of Biology, Division of Natural and Exact SciencesUniversity of GuanajuatoGuanajuato, Gto.Mexico
  2. 2.Institute of Chemical-Biological Research, Universitary CityMichoacán UniversityMoreliaMexico
  3. 3.Michoacán State Public Health LaboratoryMoreliaMexico
  4. 4.Department of Fungal Strain TechnologyBagsvaerdDenmark

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