Abstract
A transcriptional comparison of wild type and a secondary metabolite deficient Trichoderma virens mutant resulted in the identification of six genes similar to those involved in secondary metabolism in other fungi, including four cytochrome P450 genes, one O-methyl transferase and one terpene cylase. Four of the genes (three cytochrome P450s and the cyclase) are located as a cluster. Transcript levels of three of the P450 genes, the O-methyl transferase and the terpene cyclase were measured. These genes are underexpressed in the mutant, which lacks the major secondary metabolites produced by this strain, viridin and viridiol. Expression levels of clones from the differential library with similarity to fungal trehalose synthase and a hydrophobin were also underexpressed in the mutant, while a heat shock protein hsp98 homolog was not. Based on the gene expression pattern and associated secondary metabolite profile, along with similarity to other secondary metabolism pathways in related fungi, we predict that the cluster is associated with the production of a terpene. The terpene could be viridin. This is the first report on cloning of secondary metabolism related genes from T. virens, and of their organization in a cluster, in this biocontrol fungus.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Altschul SF, Gish W, Miller W, Myers EW, Lipman DJ (1990) Basic local alignment search tool. J Mol Biol 215:403–410
Bateman A, Coin L, Durbin R, Finn RD, Hollich V, Griffiths-Jones S, Khanna A, Marshall M, Moxon S, Sonnhammer EL, Studholme DJ, Yeats C, Eddy SR (2004) The Pfam protein families database. Nucleic Acids Res 32:D138–D141
Brown DW, Yu JH, Kelkar HS, Fernandes M, Nesbitt TC, Keller NP, Adams TH, Leonard TJ (1996) Twenty-five coregulated transcripts define a sterigmatocystin gene cluster in Aspergillus nidulans. Proc Natl Acad Sci USA 93:1418–1422
Brown DW, Dyer RB, McCormick SP, Kendra DF, Plattner RD (2004) Functional demarcation of the Fusarium core trichothecene gene cluster. Fungal Genet Biol 41:454–462
Calvo A, Wilson R, Bok J, Keller NP (2002) Relationship between secondary metabolism and fungal development. Microbiol Mol Biol Rev 66(3):447–459
Flaherty JE, Woloshuk CP (2004) Regulation of fumonisin biosynthesis in Fusarium verticillioides by a zinc binuclear cluster-type gene, ZFR1. Appl Environ Microbiol 70:2653–2659
Hanson JR (1995) The viridin family of steroidal antibiotics. Nat Prod Rep 12:381–384
Hedden P, Phillips AL, Rojas MC, Carrera E, Tudzynski B (2001) Gibberellin biosynthesis in plants and fungi: a case for convergent evolution? J Plant Growth Regul 20:319–331
Howell CR, Stipanovic RD (1984) Phytotoxicity to crop plants and herbicidal effects on weeds of viridiol produced by Gliocladium virens. Phytopathology 74:1346–1349
Howell CR, Stipanovic RD (1995) Mechanisms in the biocontrol of Rhizoctonia solani-induced cotton seedling disease by Gliocladium virens: antibiotics. Biol Control 85:469–472
Howell CR, Stipanovic RD, Lumsden R (1993) Antibiotic production by strains of Gliocladium virens and its relation to the biocontrol of cotton seedling diseases. Biocontrol Sci Technol 3:435–441
Ihle NT, Williams R, Chow S, Chew W, Berggren MI, Paine-Murrieta G, Minion DJ, Halter RJ, Wipf P, Abraham R, Kirkpatrick L, Powis G (2004) Molecular pharmacology and antitumor activity of PX-866, a novel inhibitor of phosphoinositide-3-kinase signaling. Mol Cancer Ther 3:763–772
Jones RW, Hancock JG (1987) Conversion of viridin to viridiol by viridin-producing fungi. Can J Microbiol 33:963–966
Jones J, Dunsmuir P, Bedbrook J (1985) High-level expression of induced chimeric genes in regenerated transformed plants. EMBO J 4:2411–2418
Keller NP, Turner G, Bennett JW (2005) Fungal secondary metabolism—from biochemistry to genomics. Nat Rev Microbiol 3:937–947
Kim S, Ahn IP, Lee YH (2001) Analysis of genes expressed during rice–Magnaporthe grisea interactions. Mol Plant Microbe Interact 14:1340–1346
Kimura M, Tokai T, O’Donnell K, Ward TJ, Fujimura M, Hamamoto H, Shibata T, Yamaguchi I (2003) The trichothecene biosynthesis gene cluster of Fusarium graminearum F15 contains a limited number of essential pathway genes and expressed non-essential genes. FEBS Lett 539:105–110
Machida M, Asai K, Sano M, Tanaka T, Kumagai T, Terai G, Kusumoto K, Arima T, Akita O, Kashiwagi Y, Abe K, Gomi K, Horiuchi H, Kitamoto K, Kobayashi T, Takeuchi M, Denning DW, Galagan JE, Nierman WC, Yu J, Archer DB, Bennett JW, Bhatnagar D, Cleveland TE, Fedorova ND, Gotoh O, Horikawa H, Hosoyama A, Ichinomiya M, Igarashi R, Iwashita K, Juvvadi PR, Kato M, Kato Y, Kin T, Kokubun A, Maeda H, Maeyama N, Maruyama J, Nagasaki H, Nakajima T, Oda K, Okada K, Paulsen I, Sakamoto K, Sawano T, Takahashi M, Takase K, Terabayashi Y, Wortman JR, Yamada O, Yamagata Y, Anazawa H, Hata Y, Koide Y, Komori T, Koyama Y, Minetoki T, Suharnan S, Tanaka A, Isono K, Kuhara S, Ogasawara N, Kikuchi H (2005) Genome sequencing and analysis of Aspergillus oryzae. Nature 438:1157–1161
Malonek S, Bomke C, Bornberg-Bauer E, Rojas MC, Hedden P, Hopkins P, Tudzynski B (2005a) Distribution of gibberellin biosynthetic genes and gibberellin production in the Gibberella fujikuroi species complex. Phytochemistry 66:1296–1311
Malonek S, Rojas MC, Hedden P, Gaskin P, Hopkins P, Tudzynski B (2005b) Functional characterization of two cytochrome P450 monooxygenase genes, P450-1 and P450-4, of the gibberellic acid gene cluster in Fusarium proliferatum (Gibberella fujikuroi MP-D). Appl Environ Microbiol 71:1462–1472
Marchler-Bauer A, Bryant SH (2004) CD-Search: protein domain annotations on the fly. Nucleic Acids Res 32:W327–331
McCormick SP, Harris LJ, Alexander NJ, Ouellet T, Saparno A, Allard S, Desjardins AE (2004) Tri1 in Fusarium graminearum encodes a P450 oxygenase. Appl Environ Microbiol 70:2044–2051
Mukherjee PK, Mukhopadhyay AN (1993) Induction of stable mutants of Gliocladium virens by gamma-irradiation. Indian Phytopathol 46:393–397
Mukherjee PK, Shreshtha SM, Mukhopadhyay AN (1993) Baiting with Sclerotium rolfsii for selective isolation of Gliocladium virens from natural soil. Biocontrol Sci Technol 3:101–104
Mukherjee PK, Latha J, Hadar R, Horwitz BA (2003) TmkA, a mitogen activated protein kinase of Trichoderma virens, is involved in biocontrol properties and repression of conidiation in the dark. Eukaryot Cell 2:46–455
Mukherjee PK, Latha J, Hadar R, Horwitz BA (2004) Role of two G-protein alpha subunits, TgaA and TgaB, in the antagonism of Trichoderma virens against plant pathogens. Appl Environ Microbiol 70:542–549
Mukhopadhyay AN, Shrestha SM, Mukherjee PK (1992) Biological seed treatment for control of soil-borne plant pathogens. FAO Plant Protection Bull 40:21–30
Rynkiewicz MJ, Cane DE, Christianson DW (2001) Structure of trichodiene synthase from Fusarium sporotrichioides provides mechanistic inferences on the terpene cyclization cascade. Proc Natl Acad Sci USA 98:13543–13548
Roberts DW (1992) Cloning and regulatory analysis of starvation-stress gene, ssgA, encoding a hydrophobin-like protein from the entomopathogenic fungus, Metarhizium anisopliae. Gene 120:119–124
Sambrook J, Fritsch EF, Maniatis T (1989) Molecular cloning: a laboratory manual. 2nd edn. Cold Spring Harbor, NY
Sivasithamparam K, Ghisalberti E (1998) Secondary metabolism in Trichoderma and Gliocladium. In: Kubicek CP, Harman GE (eds) Trichoderma and Gliocladium basic biology, taxonomy and genetics, vol 1 Taylor Francis, London, pp 139–191
St Leger RJ, Staples RC, St Leger RJ, Staples RC, Roberts DW (1992) Cloning and regulatory analysis of starvation-stress gene, ssgA, encoding a hydrophobin-like protein from the entomopathogenic fungus, Metarhizium anisopliae. Gene 120:119–124
Viterbo A, Harel M, Horwitz BA, Chet I, Mukherjee PK (2005) Trichoderma mitogen-activated protein kinase signaling is involved in induction of plant systemic resistance. Appl Environ Microbiol 71:6241–6246
Wipf P, Minion DJ, Halter RJ, Berggren MI, Ho CB, Chiang GG, Kirkpatrick L, Abraham R, Powis G (2004) Synthesis and biological evaluation of synthetic viridins derived from C (20)-heteroalkylation of the steroidal PI-3-kinase inhibitor wortmannin. Org Biomol Chem 2:1911–1920
Ying SH, Feng MG (2004) Relationship between thermotolerance and hydrophobin-like proteins in aerial conidia of Beauveria bassiana and Paecilomyces fumosoroseus as fungal biocontrol agents. J Appl Microbiol 97:323–331
Yu JH, Keller NP (2005) Regulation of secondary metabolism in filamentous fungi. Annu Rev Phytopathol 43:437–458
Acknowledgments
Thanks are due to the Department of Biotechnology, Government of India, and the Ministry of Science, Israel, for an India-Israel Research Grant, and to Dr. CR Howell for help in confirming our strain as a “P” strain and also in confirming the identity of the secondary metabolites. We are grateful to Dr. Oded Béjà for help in assembling the cosmid sequence. PKM thanks Dr. SP Kale, Head, PRSS, NABTD and Dr. SF D’Souza, Head, NABTD for encouragement and support.
Author information
Authors and Affiliations
Corresponding author
Additional information
Communicated by U. Kück
Rights and permissions
About this article
Cite this article
Mukherjee, M., Horwitz, B.A., Sherkhane, P.D. et al. A secondary metabolite biosynthesis cluster in Trichoderma virens: evidence from analysis of genes underexpressed in a mutant defective in morphogenesis and antibiotic production. Curr Genet 50, 193–202 (2006). https://doi.org/10.1007/s00294-006-0075-0
Received:
Revised:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s00294-006-0075-0