Skip to main content

Advertisement

SpringerLink
Log in

Real-time quantitative analysis of carbon catabolite derepression of cellulolytic genes expressed in the basidiomycete Phanerochaete chrysosporium

  • Applied Genetics and Molecular Biotechnology
  • Published:
Applied Microbiology and Biotechnology Aims and scope Submit manuscript

Abstract

Production of cellulolytic enzymes, such as cellobiohydrolases (CBH) and cellobiose dehydrogenase (CDH), by the basidiomycete Phanerochaete chrysosporium is significantly repressed in glucose-containing media; this is known as carbon catabolite repression. We have analyzed the glucose concentration dependence of transcript numbers of the cellulolytic genes (cel6A, cel7D, and cdh) and β-glucosidase gene (bgl3A) by means of real-time quantitative reverse transcriptase polymerase chain reaction to investigate the roll of carbon catabolite derepression in these gene expression. When the mycelium of P. chrysosporium grown in glucose culture was transferred to media containing various concentrations of glucose (0–5,000 μM), the expression levels of cel6A, cel7D, and cdh were drastically influenced by glucose, whereas no significant change was observed in bgl3A. The numbers of transcripts of cel6A, cel7D, and cdh increased exponentially during incubation for 6 h in the culture without glucose, and the rates of increase were 2.1 times per hour for cel6A transcripts and 2.7 times per hour for cel7D transcripts. Moreover, derepression of cel6A and cel7D was delayed (by 1.6 and 0.6 h, respectively) when the culture contained 50 μM glucose compared with that in the absence of glucose, suggesting that the promoter activities of cel7D and cel6A are distinct under conditions of carbon catabolite derepression.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5

Similar content being viewed by others

References

  • Aro N, Pakula T, Penttila M (2005) Transcriptional regulation of plant cell wall degradation by filamentous fungi. FEMS Microbiol Rev 29:719–739

    Article  CAS  PubMed  Google Scholar 

  • Ayers AR, Ayers SB, Eriksson KE (1978) Cellobiose oxidase, purification and partial characterization of a hemoprotein from Sporotrichum pulverulentum. Eur J Biochem 90:171–181

    Article  CAS  PubMed  Google Scholar 

  • Bao W, Usha SN, Renganathan V (1993) Purification and characterization of cellobiose dehydrogenase, a novel extracellular hemoflavoenzyme from the white-rot fungus Phanerochaete chrysosporium. Arch Biochem Biophys 300:705–713

    Article  CAS  PubMed  Google Scholar 

  • Broda P, Birch PR, Brooks PR, Sims PF (1995) PCR-mediated analysis of lignocellulolytic gene transcription by Phanerochaete chrysosporium: substrate-dependent differential expression within gene families. Appl Environ Microbiol 61:2358–2364

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Canevascini G, Coudray M-R, Rey J-P, Southgate RJG, Meier H (1979) Induction and catabolite repression of cellulase synthesis in the thermophilic fungus Sporotrichum thermophile. J Gen Microbiol 110:291–303

    Article  CAS  Google Scholar 

  • Covert SF, Vanden Wymelenberg A, Cullen D (1992) Structure, organization, and transcription of a cellobiohydrolase gene cluster from Phanerochaete chrysosporium. Appl Environ Microbiol 58:2168–2175

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Cubero B, Scazzocchio C (1994) Two different, adjacent and divergent zinc finger binding sites are necessary for CREA-mediated carbon catabolite repression in the proline gene cluster of Aspergillus nidulans. EMBO J 13:407–415

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Deshpande V, Eriksson KE, Pettersson B (1978) Production, purification and partial characterization of 1,4-beta-glucosidase enzymes from Sporotrichum pulverulentum. Eur J Biochem 90:191–198

    Article  CAS  PubMed  Google Scholar 

  • Dowzer CE, Kelly JM (1989) Cloning of the creA gene from Aspergillus nidulans: a gene involved in carbon catabolite repression. Curr Genet 15:457–459

    Article  CAS  PubMed  Google Scholar 

  • Dowzer CE, Kelly JM (1991) Analysis of the creA gene, a regulator of carbon catabolite repression in Aspergillus nidulans. Mol Cell Biol 11:5701–5709

    CAS  PubMed  PubMed Central  Google Scholar 

  • Eberhart BM, Beck RS, Goolsby KM (1977) Cellulase of Neurospora crassa. J Bacteriol 130:181–186

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Eriksson KE (1978) Enzyme mechanisms involved in cellulose hydrolysis by the rot fungus Sporotrichum pulverulentum. Biotechnol Bioeng 20:317–332

    Article  CAS  Google Scholar 

  • Henriksson G, Sild V, Szabo IJ, Pettersson G, Johansson G (1998) Substrate specificity of cellobiose dehydrogenase from Phanerochaete chrysosporium. Biochim Biophys Acta 1383:48–54

    Article  CAS  PubMed  Google Scholar 

  • Igarashi K, Tani T, Kawai R, Samejima M (2003) Family 3 beta-glucosidase from cellulose-degrading culture of the white-rot fungus Phanerochaete chrysosporium is a glucan 1,3-beta-glucosidase. J Biosci Bioeng 95:572–576

    Article  CAS  PubMed  Google Scholar 

  • Ilmen M, Thrane C, Penttila M (1996a) The glucose repressor gene cre1 of Trichoderma: isolation and expression of a full-length and a truncated mutant form. Mol Gen Genet 251:451–460

    CAS  PubMed  Google Scholar 

  • Ilmen M, Onnela ML, Klemsdal S, Keranen S, Penttila M (1996b) Functional analysis of the cellobiohydrolase I promoter of the filamentous fungus Trichoderma reesei. Mol Gen Genet 253:303–314

    CAS  PubMed  Google Scholar 

  • Ilmen M, Saloheimo A, Onnela ML, Penttila ME (1997) Regulation of cellulase gene expression in the filamentous fungus Trichoderma reesei. Appl Environ Microbiol 63:1298–1306

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Johnsrud SC, Eriksson KE (1985) Cross-breeding of selected and mutated homokaryotic strains of Phanerochaete chrysosporium K-3—new cellulase deficient strains with increased ability to degrade lignin. Appl Microbiol Biotechnol 21:320–327

    Article  CAS  Google Scholar 

  • Kremer SM, Wood PM (1992) Evidence that cellobiose oxidase from Phanerochaete chrysosporium is primarily an Fe(III) reductase. Kinetic comparison with neutrophil NADPH oxidase and yeast flavocytochrome b2. Eur J Biochem 205:133–138

    Article  CAS  PubMed  Google Scholar 

  • Kurasawa T, Yachi M, Suto M, Kamagata Y, Takao S, Tomita F (1992) Induction of cellulase by gentiobiose and its sulfur-containing analog in Penicillium purpurogenum. Appl Environ Microbiol 58:106–110

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Lamar RT, Schoenike B, Vanden Wymelenberg A, Stewart P, Dietrich DM, Cullen D (1995) Quantitation of fungal mRNAs in complex substrates by reverse transcription PCR and its application to Phanerochaete chrysosporium-colonized soil. Appl Environ Microbiol 61:2122–2126

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Li B, Nagalla SR, Renganathan V (1996) Cloning of a cDNA encoding cellobiose dehydrogenase, a hemoflavoenzyme from Phanerochaete chrysosporium. Appl Environ Microbiol 62:1329–1335

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Lymar ES, Li B, Renganathan V (1995) Purification and characterization of a cellulose-binding (beta)-glucosidase from cellulose-degrading cultures of Phanerochaete chrysosporium. Appl Environ Microbiol 61:2976–2980

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Mandels M, Parrish FW, Reese ET (1962) Sophorose as an inducer of cellulase in Trichoderma viride. J Bacteriol 83:400–408

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Meijer MM, Boonstra J, Verkleij AJ, Verrips CT (1998) Glucose repression in Saccharomyces cerevisiae is related to the glucose concentration rather than the glucose flux. J Biol Chem 273:24102–24107

    Article  CAS  PubMed  Google Scholar 

  • Ohnishi Y, Nagase M, Ichiyanagi T, Kitamoto Y, Aimi T (2007) Transcriptional regulation of two cellobiohydrolase encoding genes (cel1 and cel2) from the wood-degrading basidiomycete Polyporus arcularius. Appl Microbiol Biotechnol 76:1069–78

    Article  CAS  PubMed  Google Scholar 

  • Rho D, Desrochers M, Jurasek L, Driguez H, Defaye J (1982) Induction of cellulose in Schizophyllum commune: thiocellobiose as a new inducer. J Bacteriol 149:47–53

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Schmoll M, Kubicek CP (2003) Regulation of Trichoderma cellulase formation: lessons in molecular biology from an industrial fungus. A review. Acta Microbiol Immunol Hung 50:125–145

    Article  CAS  PubMed  Google Scholar 

  • Seiboth B, Hakola S, Mach RL, Suominen PL, Kubicek CP (1997) Role of four major cellulases in triggering of cellulase gene expression by cellulose in Trichoderma reesei. J Bacteriol 179:5318–5320

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Sims P, James C, Broda P (1988) The identification, molecular cloning and characterisation of a gene from Phanerochaete chrysosporium that shows strong homology to the exo-cellobiohydrolase I gene from Trichoderma reesei. Gene 74:411–422

    Article  CAS  PubMed  Google Scholar 

  • Sims PF, Soares-Felipe MS, Wang Q, Gent ME, Tempelaars C, Broda P (1994) Differential expression of multiple exo-cellobiohydrolase I-like genes in the lignin-degrading fungus Phanerochaete chrysosporium. Mol Microbiol 12:209–216

    Article  CAS  PubMed  Google Scholar 

  • Smith MH, Gold MH (1979) Phanerochaete chrysosporium beta-glucosidases: Induction, cellular localization, and physical characterization. Appl Environ Microbiol 37:938–942

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Stapleton PC, Dobson AD (2003) Carbon repression of cellobiose dehydrogenase production in the white rot fungus Trametes versicolor is mediated at the level of gene transcription. FEMS Microbiol Lett 221:167–172

    Article  CAS  PubMed  Google Scholar 

  • Strauss J, Mach RL, Zeilinger S, Hartler G, Stoffler G, Wolschek M, Kubicek CP (1995) Cre1, the carbon catabolite repressor protein from Trichoderma reesei. FEBS Lett 376:103–107

    Article  CAS  PubMed  Google Scholar 

  • Streamer M, Eriksson KE, Pettersson B (1975) Extracellular enzyme-system utilized by fungus Sporotrichum-Pulverulentum (Chrysosporium-Lignorum) for breakdown of cellulose - Functional characterization of 5 endo-1,4-beta-glucanases and one exo-1,4-beta-glucanase. Eur J Biochem 59:607–613

    Article  CAS  PubMed  Google Scholar 

  • Takashima S, Iikura H, Nakamura A, Masaki H, Uozumi T (1996) Analysis of Cre1 binding sites in the Trichoderma reesei cbh1 upstream region. FEMS Microbiol Lett 145:361–366

    Article  CAS  PubMed  Google Scholar 

  • Tempelaars CA, Birch PR, Sims PF, Broda P (1994) Isolation, characterization, and analysis of the expression of the cbhII gene of Phanerochaete chrysosporium. Appl Environ Microbiol 60:4387–4393

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Uzcategui E, Ruiz A, Montesino R, Johansson G, Pettersson G (1991) The 1,4-beta-D-glucan cellobiohydrolases from Phanerochaete-Chrysosporium. 1. A system of synergistically acting enzymes homologous to Trichoderma-Reesei. J Biotechnol 19:271–285

    Article  CAS  PubMed  Google Scholar 

  • Vallim MA, Janse BJ, Gaskell J, Pizzirani-Kleiner AA, Cullen D (1998) Phanerochaete chrysosporium cellobiohydrolase and cellobiose dehydrogenase transcripts in wood. Appl Environ Microbiol 64:1924–1928

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Vanden Wymelenberg A, Covert SF, Cullen D (1993) Identification of the gene encoding the major cellobiohydrolase of the white rot fungus Phanerochaete chrysosporium. Appl Envir Microbiol 59:3492–3494

    Article  CAS  Google Scholar 

  • Vanden Wymelenberg A, Minges P, Sabat G et al (2006) Computational analysis of the Phanerochaete chrysosporium v2.0 genome database and mass spectrometry identification of peptides in ligninolytic cultures reveal complex mixtures of secreted proteins. Fungal Genet Biol 43:343–356

    Article  CAS  PubMed  Google Scholar 

  • Yoshida M, Igarashi K, Kawai R, Aida K, Samejima M (2004) Differential transcription of beta-glucosidase and cellobiose dehydrogenase genes in cellulose degradation by the basidiomycete Phanerochaete chrysosporium. FEMS Microbiol Lett 235:177–182

    CAS  PubMed  Google Scholar 

  • Zeilinger S, Schmoll M, Pail M, Mach RL, Kubicek CP (2003) Nucleosome transactions on the Hypocrea jecorina (Trichoderma reesei) cellulase promoter cbh2 associated with cellulase induction. Mol Genet Genomics 270:46–55

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgments

The authors are grateful to Professor K. Aida, Associate Professor T. Kaneko and Mr. S. Ishii for technical assistance on real-time quantitative PCR. This research was supported by a Grant-in-Aid for Scientific Research to M.S. (no. 17380102) from the Japanese Ministry of Education, Culture, Sports, and Technology, and a High Efficiency Bioenergy Conversion Project to M.S. (no. 07003004-0) from New Energy and Industrial Technology Development Organization (NEDO).

Author information

Authors and Affiliations

  1. Department of Biomaterials Sciences, Graduate School of Agricultural and Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo, 113-8657, Japan

    Hitoshi Suzuki, Kiyohiko Igarashi & Masahiro Samejima

Authors
  1. Hitoshi Suzuki
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Kiyohiko Igarashi
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Masahiro Samejima
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Masahiro Samejima.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Suzuki, H., Igarashi, K. & Samejima, M. Real-time quantitative analysis of carbon catabolite derepression of cellulolytic genes expressed in the basidiomycete Phanerochaete chrysosporium . Appl Microbiol Biotechnol 80, 99–106 (2008). https://doi.org/10.1007/s00253-008-1539-6

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00253-008-1539-6

Keywords

Search

Navigation