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Current Genetics

, Volume 55, Issue 2, pp 175–184 | Cite as

Carotenoids and carotenogenic genes in Podospora anserina: engineering of the carotenoid composition extends the life span of the mycelium

  • Ingmar Strobel
  • Jürgen Breitenbach
  • Christian Q. Scheckhuber
  • Heinz D. Osiewacz
  • Gerhard SandmannEmail author
Research Article

Abstract

Carotenoids have been identified in the fungus Podospora anserina and a parallel pathway to neurosporene and β-carotene was established. Three genes for the β-carotene branch have been cloned and their function elucidated. They correspond to the al-1, al-2 and al-3 genes from Neurospora crassa. They were individually and in combinations over-expressed in P. anserina in order to modify the carotenoid composition qualitatively and quantitatively. In the resulting transformants, carotenoid synthesis was up to eightfold increased and several intermediates of the pathway together with special cyclic carotenoids, β-zeacarotene and 7,8-dihydro-β-carotene, accumulated. All transformants with an over-expressed al-2 gene (encoding a phytoene synthase and a lycopene cyclase) displayed up to 31% prolonged life span.

Keywords

Ageing protection al Genes Carotenoid biosynthesis Metabolic engineering Neurosporaxanthin 

Notes

Acknowledgments

This work was supported by the Förderfond der Goethe Universität Frankfurt and grant FP7-KBBE-2007-207948 by the European Commission to G. Sandmann and LSHM-CT-2004-512020 to H. D. Osiewacz.

References

  1. Aasen AJ, Jensen SL (1965) Fungal carotenoids. II. The structure of the carotenoid acid neurosporaxanthin. Acta Chem Scand 19:1843–1853PubMedCrossRefGoogle Scholar
  2. Arpin N, Liaaen-Jensen S (1967a) Recherches chimotaxinomiques sur les champignons Fungal carotenoids. III-Nouveaux carotenoides, notamment sous forme d′esters tertiaires, isoles de Plectania coccinea (Scop. Ex Fr) Fuck. Phytochem 6:995–1005CrossRefGoogle Scholar
  3. Arpin N, Liaaen-Jensen S (1967b) Recherches chimiotaxinomiques sur le champignons. Sur la présence de l′ester méthylique de la torularhodine chez Cookeina sulcipes. C.R. Acad Sci Paris 265:1083–1085Google Scholar
  4. Arpin N, Kjϕsen H, Francis GW, Liaaen-Jensen S (1973) The structure of aleuriaxanthin. Phytochem 12:2751–2758CrossRefGoogle Scholar
  5. Avalos J, Mackenzie A, Nelki DS, Bramley PM (1988) Terpenoid biosynthesis in cell extracts of wild-type and mutant strains of Gibberella fujikuroi. Biochim Biophys Acta 966:257–265Google Scholar
  6. Averbeck NB, Borghouts C, Hamann A, Specke V, Osiewacz HD (2001) Molecular control of copper homeostasis in filamentous fungi: increased expression of a metallothionein gene during aging of Podospora anserina. Mol Gen Genet 264:604–612PubMedCrossRefGoogle Scholar
  7. Bahadorani S, Bahadorani P, Phillips JP, Hilliker AJ (2008) The effects of vitamin supplementation on Drosophila life span under normoxia and under oxidative stress. J Gerontol A Biol Sci Med Sci 63:35–42PubMedGoogle Scholar
  8. Blanc PL, Tuveson RW, Sargent ML (1976) Inactivation of carotenoid-producing and albino strains of Neurospora crassa by visible light, black light and ultraviolet radiation. J Bacteriol 125:616–625PubMedGoogle Scholar
  9. Borovkov AY, Rivkin MI (1997) XcmI-Containing vector for direct cloning of PCR products. Biotechniques 22:812–814PubMedGoogle Scholar
  10. Breitenbach J, Braun G, Steiger S, Sandmann G (2001) Chromatographic performance on a C30-bonded stationary phase of mono hydroxycarotenoids with variable chain length or degree of desaturation and of lycopene isomers synthesized by different carotene desaturases. J Chromatogr A 936:59–69PubMedCrossRefGoogle Scholar
  11. Britton G (1995) Structure and properties of carotenoids in relation to function. FASEB J 9:1551–1558PubMedGoogle Scholar
  12. Buzzini P, Inoocenti M, Turchetti B, Libkind D, van Broock M, Mulinacci N (2007) Carotenoid profiles of yeasts belonging to the genera Rhodotorula, Rhodosporidium, Sporobolomyces and Sporidiobolus. Can J Microbiol 53:1024–1031PubMedCrossRefGoogle Scholar
  13. Edge R, McGarvey DJ, Truscott TG (1997) The carotenoids as anti-oxidants—a review. J Photochem Photobiol B 41:189–200PubMedCrossRefGoogle Scholar
  14. Espagne E, Lespinet O, Malagnac F, Da Silva C, Jaillon O, Porcel BM, Couloux A, Aury M, Segurens B, Poulain J, Anthouard V, Grossetete S, Khalili H, Coppin E, Dequard-Chablat M, Picard M, Contamine V, Arnaise S, Bourdais A, Berteaux-Lecellier V, Gautheret D, de Vries RP, Battaglia E, Coutinho PM, Danchin EGJ, Henrissat B, Khoury REL, Sainsard-Chanet A, Boivin A, Pinan-Lucarre B, Sellem CH, Debuchy R, Wincker P, Weissenbach J, Silar P (2008) The genome sequence of the model ascomycete fungus Podospora anserina. Gen Biol 9:R77-1–R77-22CrossRefGoogle Scholar
  15. Esser K (1974) Podospora anserina. In: King RC (ed) Handbook of genetics. Plenum Press, New York, pp 531–551Google Scholar
  16. Estrada AF, Youssar L, Scherzinger D, Al-Babili S, Avalos J (2008) The ylo-1 gene encodes an aldehyde dehydrogenase responsible for the last reaction in the Neurospora carotenoid pathway. Mol Microbiol 69:1207–1220PubMedCrossRefGoogle Scholar
  17. Goodwin TW (1980) The biochemistry of the carotenoids. Vol. I Plants, vol 2. Chapman and Hall, LondonGoogle Scholar
  18. Harman D (1956) A theory based on free radical and radiation chemistry. J Gerontol 11:298–300PubMedGoogle Scholar
  19. Harman D (1972) The biologic clock: the mitochondria? J Am Geriatr Soc 20:145–147PubMedGoogle Scholar
  20. Harman D (1992) Free radical theory of aging. Mutat Res 275:257–266PubMedGoogle Scholar
  21. Hausmann A, Sandmann G (2000) A single five-step desaturase is involved in the carotenoid biosynthesis pathway to beta-carotene and torulene in Neurospora crassa. Fungal Genet Biol 30:147–153PubMedCrossRefGoogle Scholar
  22. Herz S, Weber RWS, Anke H, Mucci A, Davoli P (2007) Intermediates in the oxidative pathway from torulene to torularhodin in the red yeasts Cystofilobasidium infirmominiatum and C. capitatum (Heterobasidiomycetes, Fungi). Phytochem 68:2503–2511CrossRefGoogle Scholar
  23. Lorin S, Dufour E, Sainsard-Chanet A (2006) Mitochondrial metabolism and aging in the filamentous fungus Podospora anserina. Biochim Biophys Acta 1757:604–610PubMedCrossRefGoogle Scholar
  24. Misawa N, Nakagawa M, Kobayashi K, Yamano S, Izawa Y, Nakamura K, Harashima K (1990) Elucidation of the Erwinia uredovora carotenoid biosynthetic pathway by functional analysis of gene products expressed in Escherichia coli. J Bacteriol 172:6704–6712PubMedGoogle Scholar
  25. Misawa N, Satomi Y, Kondo K, Yokoyama A, Kajiwara S, Saito T, Ohtani T, Miki W (1995) Structure and functional analysis of a marine bacterial carotenoid biosynthesis gene cluster and astaxanthin biosynthetic pathway proposed at the gene level. J Bacteriol 177:6575–6584PubMedGoogle Scholar
  26. Miura Y, Kondo K, Saito T, Shimada H, Fraser PD, Misawa N (1998) Production of the carotenoids lycopene, beta-carotene, and astaxanthin in the food yeast Candida utilis. Appl Environ Microbiol 64:1226–1229PubMedGoogle Scholar
  27. Ojima K, Breitenbach J, Visser H, Setoguchi Y, Tabata K, Hoshino T, van den Berg J, Sandmann G (2006) Cloning of the astaxanthin synthase gene from Xanthophyllomyces dendrorhous (Phaffia rhodozyma) and its assignment as a beta-carotene 3-hydroxylase/4-ketolase. Mol Genet Genomics 275:148–158PubMedCrossRefGoogle Scholar
  28. Osiewacz HD (2002) Mitochondrial functions and aging. Gene 286:65–71PubMedCrossRefGoogle Scholar
  29. Osiewacz HD, Skaletz A, Esser K (1991) Integrative transformation of the ascomycete Podospora anserina: identification of the mating-type locus on chromosome VII of electrophoretically separated chromosomes. Appl Microbiol Biotechnol 35:38–45PubMedCrossRefGoogle Scholar
  30. Rau W, Schrott EL (1987) Blue light control of pigment biosynthesis. In: Senger H (ed) Blue light responses. l, CRC Press, pp 43–63Google Scholar
  31. Saelices L, Youssar L, Holdermann I, Al-Babili S, Avalos J (2007) Identification of the gene responsible for torulene cleavage in the Neurospora carotenoid pathway. Mol Genet Genomics 278:527–537PubMedCrossRefGoogle Scholar
  32. Sander LC, Sharpless KE, Craft NE, Wise SA (1994) Development of engineered stationary phases for the separation of carotenoid isomers. Anal Chem 66:1667–1674PubMedCrossRefGoogle Scholar
  33. Sandmann G (2002) Combinatorial biosynthesis of carotenoids in a heterologous host: a powerful approach for the biosynthesis of novel structures. Chembiochem 3:629–635PubMedCrossRefGoogle Scholar
  34. Sandmann G, Misawa N (2002) Fungal carotenoids. In: Osiewacz HD (ed) The Mycota X. Industrial applications. Springer, Berlin, pp 247–262Google Scholar
  35. Sandmann G, Zhu C, Krubasik P, Fraser PD (2006) The biotechnological potential of the al-2 gene from Neurospra crassa for the production of monocyclic keto hydroxy carotenoids. Biochim Biophys Acta 1761:1085–1092PubMedGoogle Scholar
  36. Sandmann G, Takaichi S, Fraser PD (2008) C35-apocarotenoids in the yellow mutant Neurospora crassa YLO. Phytochem 69:2885–2890CrossRefGoogle Scholar
  37. Satomi Y, Misawa N, Maoka T, Nishino H (2004) Production of phytoene, a carotenoid, and induction of connexin 26 in transgenic mice carrying the phytoene synthase gene crtB. Biochem Biophys Res Commun 320:398–401PubMedCrossRefGoogle Scholar
  38. Scheckhuber CQ, Osiewacz HD (2008) Podospora anserina: a model organism to study mechanisms of healthy ageing. Mol Genet Genomics 280:365–374PubMedCrossRefGoogle Scholar
  39. Schmidhauser TJ, Lauter FR, Russo VE, Yanofsky C (1990) Cloning, sequence, and photoregulation of al-1, a carotenoid biosynthetic gene of Neurospora crassa. Mol Cell Biol 10:5064–5070PubMedGoogle Scholar
  40. Schmidhauser TJ, Lauter FR, Schumacher M, Zhou W, Russo VE, Yanofsky C (1994) Characterization of al-2, the phytoene synthase gene of Neurospora crassa. Cloning, sequence analysis, and photoregulation. J Biol Chem 269:12060–12066PubMedGoogle Scholar
  41. Schrantz JP, Lemoine Y (1995) Carotenoid composition of mycelium and apothecia in the discomycete Scutellinia umbrarum. Photochem 40:33–35CrossRefGoogle Scholar
  42. Siefermann-Harms D (1987) The light-harvesting and protective functions of carotenoids in photosynthetic membranes. Physiol Plant 69:661–668CrossRefGoogle Scholar
  43. Stumpferl SW, Stephan O, Osiewacz HD (2004) Impact of a disruption of a pathway delivering copper to mitochondria on Podospora anserina metabolism and life span. Eukaryot Cell 3:200–211PubMedCrossRefGoogle Scholar
  44. Thewes S, Prado-Cabrero A, Prado MM, Tudzynski B, Avalos J (2005) Characterization of a gene in the car cluster of Fusarium fujikuroi that codes for a protein of the carotenoid oxygenase family. Mol Genet Genomics 274:217–228PubMedCrossRefGoogle Scholar
  45. Verdoes CJ, Sandmann G, Visser H, Diaz M, van Mossel M, van Ooyen AJJ (2003) Metabolic engineering of the carotenoid biosynthetic pathway in the yeast Xanthophyllomyces dendrorhous (Phaffia rhodozyma). Appl Environ Microbiol 69:3728–3738PubMedCrossRefGoogle Scholar
  46. Verwaal R, Wang J, Meijnen JP, Visser H, Sandmann G, van den Berg JA, van Ooyen AJ (2007) High level production of beta-carotene in Saccharomyces cerevisiae by successive transformation with carotenogenic genes from Xanthophyllomyces dendrorhous. Appl Environ Microbiol 73:4342–4350PubMedCrossRefGoogle Scholar
  47. Will OH, Newland NA, Reppe CR (1984) The photosensitivity of pigmented and non-pigmented strains of Ustilago violacea. Curr Microbiol 10:295–302CrossRefGoogle Scholar

Copyright information

© Springer-Verlag 2009

Authors and Affiliations

  • Ingmar Strobel
    • 2
  • Jürgen Breitenbach
    • 1
  • Christian Q. Scheckhuber
    • 2
  • Heinz D. Osiewacz
    • 2
  • Gerhard Sandmann
    • 1
    Email author
  1. 1.Biosynthesis Group, Department of Biological Sciences, Institute of Molecular BiosciencesJ. W. Goethe UniversitätFrankfurtGermany
  2. 2.Molecular Developmental Biology, Department of Biological Sciences and Cluster of Excellence Macromolecular Complexes, Institute of Molecular BiosciencesJ. W. Goethe UniversityFrankfurtGermany

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