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Title: Tracking the roots of cellulase hyperproduction by the fungus Trichoderma reesei using massively parallel DNA sequencing

Abstract

Trichoderma reesei (teleomorph Hypocrea jecorina) is the main industrial source of cellulases and hemicellulases harnessed for the hydrolysis of biomass to simple sugars, which can then be converted to biofuels, such as ethanol, and other chemicals. The highly productive strains in use today were generated by classical mutagenesis. To learn how cellulase production was improved by these techniques, we performed massively parallel sequencing to identify mutations in the genomes of two hyperproducing strains (NG14, and its direct improved descendant, RUT C30). We detected a surprisingly high number of mutagenic events: 223 single nucleotides variants, 15 small deletions or insertions and 18 larger deletions leading to the loss of more than 100 kb of genomic DNA. From these events we report previously undocumented non-synonymous mutations in 43 genes that are mainly involved in nuclear transport, mRNA stability, transcription, secretion/vacuolar targeting, and metabolism. This homogeneity of functional categories suggests that multiple changes are necessary to improve cellulase production and not simply a few clear-cut mutagenic events. Phenotype microarrays show that some of these mutations result in strong changes in the carbon assimilation pattern of the two mutants with respect to the wild type strain QM6a. Our analysis provides the first genome-wide insightsmore » into the changes induced by classical mutagenesis in a filamentous fungus, and suggests new areas for the generation of enhanced T. reesei strains for industrial applications such as biofuel production.« less

Authors:
; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ;
Publication Date:
Research Org.:
Pacific Northwest National Lab. (PNNL), Richland, WA (United States)
Sponsoring Org.:
USDOE
OSTI Identifier:
973985
Report Number(s):
PNNL-SA-67636
Journal ID: ISSN 0027-8424; PNASA6; BM0102070; TRN: US201007%%168
DOE Contract Number:  
AC05-76RL01830
Resource Type:
Journal Article
Journal Name:
Proceedings of the National Academy of Sciences of the United States of America, 106(38):16151-16156
Additional Journal Information:
Journal Volume: 106; Journal Issue: 38; Journal ID: ISSN 0027-8424
Country of Publication:
United States
Language:
English
Subject:
09 BIOMASS FUELS; BIOFUELS; BIOMASS; CARBON; CELLULASE; DNA; DNA SEQUENCING; ETHANOL; FUNCTIONALS; GENES; HYDROLYSIS; METABOLISM; MUTAGENESIS; MUTANTS; MUTATIONS; NUCLEOTIDES; PHENOTYPE; SACCHARIDES; STRAINS; TRANSCRIPTION; TRICHODERMA VIRIDE; Trichoderma, cellulase, fungi, genomics

Citation Formats

Le Crom, Stphane, Schackwitz, Wendy, Pennacchiod, Len, Magnuson, Jon K, Culley, David E, Collett, James R, Martin, Joel X, Druzhinina, Irina S, Mathis, Hugues, Monot, Frdric, Seiboth, Bernhard, Cherry, Barbara, Rey, Michael, Berka, Randy, Kubicek, Christian P, Baker, Scott E, and Margeot, Antoine. Tracking the roots of cellulase hyperproduction by the fungus Trichoderma reesei using massively parallel DNA sequencing. United States: N. p., 2009. Web. doi:10.1073/pnas.0905848106.
Le Crom, Stphane, Schackwitz, Wendy, Pennacchiod, Len, Magnuson, Jon K, Culley, David E, Collett, James R, Martin, Joel X, Druzhinina, Irina S, Mathis, Hugues, Monot, Frdric, Seiboth, Bernhard, Cherry, Barbara, Rey, Michael, Berka, Randy, Kubicek, Christian P, Baker, Scott E, & Margeot, Antoine. Tracking the roots of cellulase hyperproduction by the fungus Trichoderma reesei using massively parallel DNA sequencing. United States. https://doi.org/10.1073/pnas.0905848106
Le Crom, Stphane, Schackwitz, Wendy, Pennacchiod, Len, Magnuson, Jon K, Culley, David E, Collett, James R, Martin, Joel X, Druzhinina, Irina S, Mathis, Hugues, Monot, Frdric, Seiboth, Bernhard, Cherry, Barbara, Rey, Michael, Berka, Randy, Kubicek, Christian P, Baker, Scott E, and Margeot, Antoine. 2009. "Tracking the roots of cellulase hyperproduction by the fungus Trichoderma reesei using massively parallel DNA sequencing". United States. https://doi.org/10.1073/pnas.0905848106.
@article{osti_973985,
title = {Tracking the roots of cellulase hyperproduction by the fungus Trichoderma reesei using massively parallel DNA sequencing},
author = {Le Crom, Stphane and Schackwitz, Wendy and Pennacchiod, Len and Magnuson, Jon K and Culley, David E and Collett, James R and Martin, Joel X and Druzhinina, Irina S and Mathis, Hugues and Monot, Frdric and Seiboth, Bernhard and Cherry, Barbara and Rey, Michael and Berka, Randy and Kubicek, Christian P and Baker, Scott E and Margeot, Antoine},
abstractNote = {Trichoderma reesei (teleomorph Hypocrea jecorina) is the main industrial source of cellulases and hemicellulases harnessed for the hydrolysis of biomass to simple sugars, which can then be converted to biofuels, such as ethanol, and other chemicals. The highly productive strains in use today were generated by classical mutagenesis. To learn how cellulase production was improved by these techniques, we performed massively parallel sequencing to identify mutations in the genomes of two hyperproducing strains (NG14, and its direct improved descendant, RUT C30). We detected a surprisingly high number of mutagenic events: 223 single nucleotides variants, 15 small deletions or insertions and 18 larger deletions leading to the loss of more than 100 kb of genomic DNA. From these events we report previously undocumented non-synonymous mutations in 43 genes that are mainly involved in nuclear transport, mRNA stability, transcription, secretion/vacuolar targeting, and metabolism. This homogeneity of functional categories suggests that multiple changes are necessary to improve cellulase production and not simply a few clear-cut mutagenic events. Phenotype microarrays show that some of these mutations result in strong changes in the carbon assimilation pattern of the two mutants with respect to the wild type strain QM6a. Our analysis provides the first genome-wide insights into the changes induced by classical mutagenesis in a filamentous fungus, and suggests new areas for the generation of enhanced T. reesei strains for industrial applications such as biofuel production.},
doi = {10.1073/pnas.0905848106},
url = {https://www.osti.gov/biblio/973985}, journal = {Proceedings of the National Academy of Sciences of the United States of America, 106(38):16151-16156},
issn = {0027-8424},
number = 38,
volume = 106,
place = {United States},
year = {Tue Sep 22 00:00:00 EDT 2009},
month = {Tue Sep 22 00:00:00 EDT 2009}
}