Abstract
As part of an effort to determine all the gene products involved in wood degradation, we have performed massively parallel pyrosequencing on an expression library from the white rot fungus Phanerochaete chrysosporium grown in shallow stationary cultures with red oak as the carbon source. Approximately 48,000 high quality sequence tags (246 bp average length) were generated. 53% of the sequence tags aligned to 4,262 P. chrysosporium gene models, and an additional 18.5% of the tags reliably aligned to the P. chrysosporium genome providing evidence for 961 putative novel fragmented gene models. Due to their role in lignocellulose degradation, the secreted proteins were focused upon. Our results show that the four enzymes required for cellulose degradation: endocellulase, exocellulase CBHI, exocellulase CBHII, and β-glucosidase are all produced. For hemicellulose degradation, not all known enzymes were produced, but endoxylanases, acetyl xylan esterases and mannosidases were detected. For lignin degradation, the role of peroxidases has been questioned; however, our results show that lignin peroxidase is highly expressed along with the H2O2 generating enzyme, alcohol oxidase. The transcriptome snapshot reveals that H2O2 generation and utilization are central in wood degradation. Our results also reveal new transcripts that encode extracellular proteins with no known function.
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References
Abbas A, Koc H, Liu F, Tien M (2005) Fungal degradation of wood: initial proteomic analysis of extracellular proteins of Phanerochaete chrysosporium grown on oak substrate. Curr Genet 47:49–56
Adler E (1977) Lignin chemistry-past, present and future. Wood Sci Technol 11:169–218
Altschul SF, Gish W, Miller W, Myers EW, Lipman DJ (1990) Basic local alignment search tool. J Mol Biol 215:403–410
Coughlan MP, Hazlewood GP (1993) β-1, 4-D-Xylan-degrading enzyme systems—biochemistry, molecular biology and applications. Biotechnol Appl Biochem 17:259–289
Cowling E, Merrill W (1966) Nitrogen in wood and its role in wood deterioration. Can J Bot 44:1539–1554
Crawford DL, Crawford RL (1980) Microbial degradation of lignin. Enzyme Microb Technol 2:11–22
Daniel G, Volc J, Filonova L, Plihal O, Kubatova E, Halada P (2007) Characteristics of Gloeophyllum trabeum alcohol oxidase, an extracellular source of H2O2 in brown rot decay of wood. Appl Environ Microbiol 73:6241–6253
Devries OMH, Fekkes MP, Wosten HAB, Wessels JGH (1993) Insoluble hydrophobin complexes in the walls of Schizophyllum commune and other filamentous fungi. Arch Microbiol 159:330–335
Emanuelsson O, Brunak S, von Heijne G, Nielsen H (2007) Locating proteins in the cell using TargetP, SignalP and related tools. Nat Protoc 2:953–971
Emrich SJ, Barbazuk WB, Li L, Schnable PS (2007) Gene discovery and annotation using LCM-454 transcriptome sequencing. Genome Res 17:69–73
Glenn JK, Gold MH (1985) Purification and characterization of an extracellular Mn(II)-dependent peroxidase from the lignin-degrading Basidiomycete, Phanerochaete chrysosporium. Arch. Biochem Biophys 242:329–341
Glenn JK, Morgan MA, Mayfield MB, Kuwahara M, Gold MH (1983) An extracellular H2O2-requiring enzyme preparation involved in lignin biodegradation by the white rot Basidiomycete Phanerochaete chrysosporium. Biochem Biophys Res Commun 114:1077–1083
Hack CJ (2004) Integrated transcriptome and proteome data: the challenges ahead. Brief Funct Genomic Proteomic 3:212–219
Hammel KE, Cullen D (2008) Role of fungal peroxidases in biological ligninolysis. Curr. Opin. Plant Biol. 11:349–355
Hammel KE, Tien M, Kalyanaraman B, Kirk TK (1985) Mechanism of oxidative Cα-Cβ cleavage of a lignin model dimer by Phanerochaete chrysosporium ligninase. Stoichiometry and involvement of free radicals. J Biol Chem 260:8348–8353
Hendrick JP, Hartl FU (1993) Molecular chaperone functions of heat-shock proteins. Annu Rev Biochem 62:349–384
Henriksson G, Ander P, Pettersson B, Pettersson G (1995) Cellobiose dehydrogenase (cellobiose oxidase) from Phanerochaete chrysosporium as a wood degrading enzyme—studies on cellulose, xylan and synthetic lignin. Appl Microbiol Biotechnol 42:790–796
Henriksson G, Johansson G, Pettersson G (2000) A critical review of cellobiose dehydrogenases. J. Biotech. 78:93–113
Huang XQ, Madan A (1999) CAP3: A DNA sequence assembly program. Genome Res 9:868–877
Johnson KG (1990) Exocellular mannanases from hemicellulolytic fungi. World J Microbiol Biotechnol 6:209–217
Keller B, Schmid J, Lamb CJ (1989a) Vascular expression of a bean cell wall glycine-rich protein-beta-glucuronidase gene fusion in transgenic tobacco. EMBO J 8:1309–1314
Keller B, Templeton MD, Lamb CJ (1989b) Specific localization of a plant cell wall glycine-rich protein in protoxylem cells of the vascular system. Proc Natl Acad Sci USA 86:1529–1533
Kersten PJ, Kirk TK (1987) Involvement of a new enzyme, glyoxal oxidase, in extracellular H2O2 production by Phanerochaete chrysosporium. J Bacteriol 169:2195–2201
Kersten PJ, Tien M, Kalyanaraman B, Kirk TK (1985) The ligninase of Phanerochaete chrysosporium generates cation radicals from methoxybenzenes. J Biol Chem 260:2609–2612
Keyser P, Kirk TK, Zeikus JG (1978) Ligninolytic enzyme system of Phanaerochaete chrysosporium: synthesized in the absence of lignin in response to nitrogen starvation. J Bacteriol 135:790–797
Kirk TK, Adler E (1970) Methoxyl deficient structural elements in lignin of sweetgum decayed by a brown-rot fungus. Acta Chem Scand 24:3379–3390
Kirk TK (1983) Degradation and conversion of lignocelluloses. Edward Arnold, London
Kirk TK, Cullen D (1998) Enzymology and molecular genetics of wood degradation by white-rot fungi. Environmentally friendly technologies for the pulp and paper lndustry. John, Hoboken
Kirk TK, Farrell RL (1987) Enzymatic combustion: the microbial degradation of lignin. Ann Rev Microbiol 41:465–505
Kirk TK, Tien M, Kersten PJ, Kalyanaraman B, Hammel KE, Farrell RL (1990) Lignin peroxidase from fungi Phanerochaete chrysosporium. Meth Enzymol 188:159–171
Kuan IC, Johnson KA, Tien M (1993) Kinetic analysis of manganese peroxidase: the reaction with manganese complexes. J Biol Chem 268:20064–20070
Margulies M, Egholm M, Altman WE, Attiya S, Bader JS, Bemben LA, Berka J, Braverman MS, Chen YJ, Chen ZT, Dewell SB, Du L, Fierro JM, Gomes XV, Godwin BC, He W, Helgesen S, Ho CH, Irzyk GP, Jando SC, Alenquer MLI, Jarvie TP, Jirage KB, Kim JB, Knight JR, Lanza JR, Leamon JH, Lefkowitz SM, Lei M, Li J, Lohman KL, Lu H, Makhijani VB, McDade KE, McKenna MP, Myers EW, Nickerson E, Nobile JR, Plant R, Puc BP, Ronan MT, Roth GT, Sarkis GJ, Simons JF, Simpson JW, Srinivasan M, Tartaro KR, Tomasz A, Vogt KA, Volkmer GA, Wang SH, Wang Y, Weiner MP, Yu PG, Begley RF, Rothberg JM (2005) Genome sequencing in microfabricated high-density picolitre reactors. Nature 437:376–380
Martinez D, Larrondo LF, Putnam N, Gelpke MDS, Huang K, Chapman J, Helfenbein KG, Ramaiya P, Detter JC, Larimer F, Coutinho PM, Henrissat B, Berka R, Cullen D, Rokhsar D (2004) Genome sequence of the lignocellulose degrading fungus Phanerochaete chrysosporium strain RP78. Nat Biotechnol 22:695–700
Mayfield MB, Godfrey BJ, Gold MH (1994) Characterization of the Mnp2 gene encoding manganese peroxidase isozyme-2 from the Basidiomycete Phanerochaete chrysosporium. Gene 142:231–235
Paszczynski A, Huynh VB, Crawford R (1985) Enzymatic activities of an extracellular, manganese-dependent peroxidase from Phanerochaete chrysosporium. FEMS Microb Lett 29:37–41
Poinar HN, Schwarz C, Qi J, Shapiro B, MacPhee RDE, Buigues B, Tikhonov A, Huson DH, Tomsho LP, Auch A, Rampp M, Miller W, Schuster SC (2006) Metagenomics to paleogenomics: Large-scale sequencing of mammoth DNA. Science 311:392–394
Quevillon E, Silventoinen V, Pillai S, Harte N, Mulder N, Apweiler R, Lopez R (2005) InterProScan: protein domains identifier. Nucleic Acids Res 33:W116–W120
Sarkanen S, Razal RA, Piccariello T, Yamamoto E, Lewis NG (1991) Lignin peroxidase: toward a clarification of its role in vivo. J Biol Chem 266:3636–3643
Sato S, Liu F, Koc H, Tien M (2007) Expression analysis of extracellular proteins from Phanerochaete chrysosporium grown on different liquid and solid substrates. Microbiology 153:3023–3033
Shary S, Kapich AN, Panisko EA, Magnuson JK, Cullen D, Hammel KE (2008) Differential expression in Phanerochaete chrysosporium of membrane-associated proteins relevant to lignin degradation. Appl Environ Microbiol 74:7252–7725
Tien M, Kirk TK (1983) Lignin-degrading enzyme from the Hymenomycete Phanerochaete chrysosporium Burds. Science 221:661–663
Tien M, Kirk TK (1988) Lignin peroxidase of Phanerochaete chrysosporium. Meth Enzymol 161:238–249
Timmel TE (1967) Recent progress in the chemistry of wood hemicelluloses. Wood Sci Technol 1:45–70
Vanden Wymelenberg A, Sabat G, Martinez D, Rajangam AS, Teeri TT, Gaskell J, Kersten PJ, Cullen D (2005) The Phanerochaete chrysosporium secretome: database predictions and initial mass spectrometry peptide identifications in cellulose-grown medium. J Biotechnol 118:17–34
Vera JC, Wheat CW, Fescemyer HW, Frilander MJ, Crawford DL, Hanski I, Marden JH (2008) Rapid transcriptome characterization for a nonmodel organism using 454 pyrosequencing. Mol Ecol 17:1636–1647
Wang SX, Hunter W, Plant A (2000) Isolation and purification of functional total RNA from woody branches and needles of Sitka and white spruce. Biotechniques 28:292–296
Wessels JGH, Devries OMH, Asgeirsdottir SA, Schuren FHJ (1991) Hydrophobin genes involved in formation of aerial hyphae and fruit bodies in Schizophyllum. Plant Cell 3:793–799
Wosten HAB, Devries OMH, Wessels JGH (1993) Interfacial self assembly of a fungal hydrophobin into a hydrophobic rodlet layer. Plant Cell 5:1567–1574
Acknowledgments
This research was supported in part by a grant from the Department of Energy (DE-FG02-87ER13690) to MT. Partially funded by Clemson Experiment Station project #SC-1700381.
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Communicated by U. Kues.
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Sato, S., Feltus, F.A., Iyer, P. et al. The first genome-level transcriptome of the wood-degrading fungus Phanerochaete chrysosporium grown on red oak. Curr Genet 55, 273–286 (2009). https://doi.org/10.1007/s00294-009-0243-0
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DOI: https://doi.org/10.1007/s00294-009-0243-0