Abstract
Symbiotic ectomycorrhizal fungi contribute to the nitrogen nutrition of their host-plants but little information is available on the molecular control of their nitrogen metabolism. We cloned and characterised genes encoding a nitrite reductase and a nitrate transporter in the ectomycorrhizal basidiomycete Hebeloma cylindrosporum. These two genes are divergently transcribed and linked to a previously cloned nitrate reductase gene, thus demonstrating that nitrate assimilation gene clusters occur in homobasidiomycetes. The nitrate transporter polypeptide (NRT2) is characterised by 12 transmembrane domains and presents both a long putative intracellular loop and a short C-terminal tail, two structural features which distinguish fungal high-affinity transporters from their plant homologues. In different wild-type genetic backgrounds, transcription of the two genes was repressed by ammonium and was strongly stimulated not only in the presence of nitrate but also in the presence of organic nitrogen sources or under nitrogen deficiency.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Altschul SF, Madden TL, Schaffer AA, Zhang J, Zhang Z, Miller W, Lipman DJ (1997) Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Res 25:3389–3402
Amaar YG, Moore MM (1998) Mapping of the nitrate-assimilation gene cluster (crnA-niiA-niaD) and characterization of the nitrite reductase gene (niiA) in the opportunistic fungal pathogen Aspergillus fumigatus. Curr Genet 33:206–215
Attwood TK, Beck ME (1994) PRINTS—a protein motif fingerprint database. Protein Eng 7:841–848
Banks GR, Shelton PA, Kanuga N, Holden DW, Spanos A (1993) The Ustilago maydis nar1 gene encoding nitrate reductase activity: sequence and transcriptional regulation. Gene 131:69–78
Brito N, Avila J, Perez MD, Gonzalez C, Siverio JM (1996) The genes YNI1 and YNR1, encoding nitrite reductase and nitrate reductase respectively in the yeast Hansenula polymorpha, are clustered and coordinately regulated. Biochem J 317:89–95
Campbell WH, Kinghorn JR (1990) Functional domains of assimilatory nitrate reductases and nitrite reductases. Trends Biochem Sci 15:315–319
Chang P-K, Ehrlich KC, Lunz JE, Bhatnagar D, Cleveland TE, Bennett JW (1996) Characterization of the Aspergillus parasiticus niaD and niiA gene cluster. Curr Genet 30:68–75
Crawford NM, Arst HNJ (1993) The molecular genetics of nitrate assimilation in fungi and plants. Annu Rev Genet 27:115–146
Debaud J-C, Gay G (1987) In vitro fruiting under controlled conditions of the ectomycorrhizal fungus Hebeloma cylindrosporum associated with Pinus pinaster. New Phytol 105:429–435
Forde BG (2000) Nitrate transporters in plants: structure, function and regulation. Biochim Biophys Acta 1465:219–235
Fraissinet-Tachet L, Baltz R, Chong J, Kauffmann S, Fritig B, Saindrenan P (1998) Two tobacco genes induced by infection, elicitor and salicylic acid encode glucosyl transferases acting on phenylpropanoids and benzoic acid derivatives, including salicylic acid. FEBS Lett 437:319–323
Gobert A, Plassard C (2002) Differential NO3-dependent patterns of NO3 uptake in Pinus pinaster, Rhizopogon roseolus and their ectomycorrhizal association. New Phytol 154:509–516
Gryta H, Debaud J-C, Marmeisse R (2000) Population dynamics of the symbiotic mushroom Hebeloma cylindrosporum: mycelial persistence and inbreeding. Heredity 84:294–302
Henikoff S, Henikoff JG (1994) Protein family classification based on searching a database of blocks. Genomics 19:97–107
Jargeat P, Gay G, Debaud JC, Marmeisse R (2000) Transcription of a nitrate reductase gene isolated from the symbiotic basidiomycete fungus Hebeloma cylindrosporum does not require induction by nitrate. Mol Gen Genet 263:948–956
Javelle A, Rodriguez-Pastrana BR, Jacob C, Botton B, Brun A, André B, Marini AM, Chalot M (2001) Molecular characterization of two ammonium transporters from the ectomycorrhizal fungus Hebeloma cylindrosporum. FEBS Lett 505:393–398
Johnstone IL, McCabe PC, Greaves P, Gurr SJ, Cole G., Brow MAD, Unkles SE, Clutterbuck AJ, Kinghorn JR, Innis MA (1990) Isolation and characterisation of the crnA-niiA-niaD gene cluster for nitrate assimilation in Aspergillus nidulans. Gene 90:181–192
Keller NP, Hohn TM (1997) Metabolic pathway gene clusters in filamentous fungi. Fungal Genet Biol 21:17–29
Kinghorn JR, Unkles SE (1994) Inorganic nitrogen assimilation: molecular aspects. In: Martinelli SD, Kinghorn JR (eds) Aspergillus: 50 years on. Elsevier, Amsterdam, pp 181–194
Marmeisse R, Jargeat P, Wagner F, Gay G, Debaud J-C (1998) Isolation and characterization of nitrate reductase deficient mutants of the ectomycorrhizal fungus Hebeloma cylindrosporum. New Phytol 140: 311–318
Martin F, Botton B (1993) Nitrogen metabolism of ectomycorrhizal fungi and ectomycorrhizas. Adv Plant Pathol 9:83–102
Martin F, Lorillou S (1997) Nitrogen acquisition and assimilation in ectomycorrhizal systems. In: Rennenberg H, Eschrich W, Ziegler H (eds) Trees—contributions to modern tree physiology. Backhuys, Leiden, pp 423–439
Marzluf GA (1997) Genetic regulation of nitrogen metabolism in the fungi. Microbiol Mol Biol Rev 61:17–32
Montanini B, Moretto N, Soragni E, Percudani R, Ottonello S (2002) A high-affinity ammonium transporter from the mycorrhizal ascomycete Tuber borchii. Fungal Genet Biol 36:22–34
Muro-Pastor MI, Gonzalez R, Strauss J, Narendja F, Scazzocchio C (1999) The GATA factor AreA is essential for chromatin remodelling in a eukaryotic bidirectional promoter. EMBO J 18:1584–1597
Nehls U, Wiese J, Güttenberger M, Hampp R (1998) Carbon allocation in ectomycorrhizas: identification and expression analysis of an Amanita muscaria monosaccharide transporter. Mol Plant-Microbe Interact 11:167–176
Pérez MD, González C, Avila J, Brito N, Siverio JM (1997) The YNT1 gene encoding the nitrate transporter in the yeast Hansenula polymorpha is clustered with genes YNI1 and YNR1 encoding nitrite reductase and nitrate reductase, and its disruption causes inability to grow in nitrate. Biochem J 321:397–403
Plassard C, Bonafos B, Touraine B (2000) Differential effects of mineral and organic N sources, and of ectomycorrhizal infection by Hebeloma cylindrosporum, on growth and N utilization in Pinus pinaster. Plant Cell Environ 23:1195–1205
Plassard C, Guérin-Laguet EA, Véry A-A, Casarin V, Thibaud J-B (2002) Local measurements of nitrate and potassium fluxes along roots of maritime pine. Effects of ectomycorrhizal symbiosis. Plant Cell Environ 25:1–10
Punt PJ, Strauss J, Smit R, Kinghorn JR, Van den Hondel CAMJJ, Scazzocchio C (1995) The intergenic region between the divergently transcribed niiA and niaD genes of Aspergillus nidulans contains multiple NIRA binding sites which act bidirectionally. Mol Cell Biol 15:5688–5699
Rambosek J and Leach J (1987) Recombinant DNA in filamentous fungi: progress and prospects, Crit Rev Biotechnol 6:357–393
Scheromm P, Plassard C, Salsac L (1990) Regulation of nitrate reductase in the ectomycorrhizal Basidiomycete Hebeloma cylindrosporum Romagn. cultured on nitrate or ammonium. New Phytol 114:441–447
Schuren FHJ (1992) Regulation of gene expression during fruit-body development in Schizophyllum commune. PhD thesis, University of Groningen, Groningen
Sequerra J, Marmeisse R, Valla G, Normand P, Capellano A, Moiroud A (1997) Taxonomic position and intraspecific variability of the nodule forming Penicillium nodositatum inferred from R.F.L.P. analysis of the ribosomal intergenic spacer and random amplified polymorphic DNA. Mycol Res 101:465–472
Smith SE, Read DJ (1997) Mycorrhizal symbiosis, 2nd edn. Academic Press, London
Thompson JD, Higgins DG, Gibson, TJ (1994) CLUSTAL W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position specific gap penalties and weight matrix choice. Nucleic Acids Res 22:4673–4680
Unkles SE, Hawker KL, Grieve C, Campbell EI, Montague P, Kinghorn JR (1991) crnA encodes a nitrate transporter in Aspergillus nidulans. Proc Natl Acad Sci 88:204–208
Unkles SE, Zhou D, Siddigi MY, Kinghorn JR, Glass ADM (2001) Apparent genetic redundancy facilitates ecological plasticity for nitrate transport. EMBO J 20 6246–6255
Van Kan JAL, Van den Ackerveken GFJM, De Wit PJGM (1991) Cloning and characterization of the cDNA of avirulence avr9 of the fungal pathogen Cladosporium fulvum, the causal agent of tomato leaf mold. Mol Plant-Microbe Interact 4:52–59
Williams RSB, Davis MA, Howlett BJ (1995) The nitrate and nitrite reductase-encoding genes of Leptosphaeria maculans are closely linked and transcribed in the same direction. Gene 158:153–154
Wipf D, Benjdia M, Tegeder M, Frommer WB (2002) Characterization of a general amino acid permease from Hebeloma cylindrosporum. FEBS Lett 528:119–124
Acknowledgements
We would like to thank Pierre Audenis for preparing the illustrations. The Phanerochaete chrysosporium data have been provided freely by the JGI for use in this publication only. This work was supported by the Programme Prioritaire Microbiologie of the INRA.
Author information
Authors and Affiliations
Corresponding author
Additional information
Communicated by U. Kück
Rights and permissions
About this article
Cite this article
Jargeat, P., Rekangalt, D., Verner, MC. et al. Characterisation and expression analysis of a nitrate transporter and nitrite reductase genes, two members of a gene cluster for nitrate assimilation from the symbiotic basidiomycete Hebeloma cylindrosporum . Curr Genet 43, 199–205 (2003). https://doi.org/10.1007/s00294-003-0387-2
Received:
Revised:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s00294-003-0387-2