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Truncation of the transcriptional repressor protein Cre1 in Trichoderma reesei Rut-C30 turns it into an activator
BACKGROUND: The filamentous fungus Trichoderma reesei (T. reesei) is a natural producer of cellulolytic and xylanolytic enzymes and is therefore industrially used. Many industries require high amounts of enzymes, in particular cellulases. Strain improvement strategies by random mutagenesis yielded t...
Autores principales: | , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
BioMed Central
2018
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6100732/ https://www.ncbi.nlm.nih.gov/pubmed/30151221 http://dx.doi.org/10.1186/s40694-018-0059-0 |
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author | Rassinger, Alice Gacek-Matthews, Agnieszka Strauss, Joseph Mach, Robert L. Mach-Aigner, Astrid R. |
author_facet | Rassinger, Alice Gacek-Matthews, Agnieszka Strauss, Joseph Mach, Robert L. Mach-Aigner, Astrid R. |
author_sort | Rassinger, Alice |
collection | PubMed |
description | BACKGROUND: The filamentous fungus Trichoderma reesei (T. reesei) is a natural producer of cellulolytic and xylanolytic enzymes and is therefore industrially used. Many industries require high amounts of enzymes, in particular cellulases. Strain improvement strategies by random mutagenesis yielded the industrial ancestor strain Rut-C30. A key property of Rut-C30 is the partial release from carbon catabolite repression caused by a truncation of the repressor Cre1 (Cre1-96). In the T. reesei wild-type strain a full cre1 deletion leads to pleiotropic effects and strong growth impairment, while the truncated cre1-96 enhances cellulolytic activity without the effect of growth deficiencies. However, it is still unclear which function Cre1-96 has in Rut-C30. RESULTS: In this study, we deleted and constitutively expressed cre1-96 in Rut-C30. We found that the presence of Cre1-96 in Rut-C30 is crucial for its cellulolytic and xylanolytic performance under inducing conditions. In the case of the constitutively expressed Cre1-96, the cellulase activity could further be improved approximately twofold. The deletion of cre1-96 led to growth deficiencies and morphological abnormalities. An in silico domain prediction revealed that Cre1-96 has all necessary properties that a classic transactivator needs. Consequently, we investigated the cellular localization of Cre1-96 by fluorescence microscopy using an eYFP-tag. Cre1-96 is localized in the fungal nuclei under both, inducing and repressing conditions. Furthermore, chromatin immunoprecipitation revealed an enrichment of Cre1-96 in the upstream regulatory region of the main transactivator of cellulases and xylanases, Xyr1. Interestingly, transcript levels of cre1-96 show the same patterns as the ones of xyr1 under inducing conditions. CONCLUSIONS: The findings suggest that the truncation turns Cre1 into an activating regulator, which primarily exerts its role by approaching the upstream regulatory region of xyr1. The conversion of repressor proteins to potential activators in other biotechnologically used filamentous fungi can be applied to increase their enzyme production capacities. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1186/s40694-018-0059-0) contains supplementary material, which is available to authorized users. |
format | Online Article Text |
id | pubmed-6100732 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2018 |
publisher | BioMed Central |
record_format | MEDLINE/PubMed |
spelling | pubmed-61007322018-08-27 Truncation of the transcriptional repressor protein Cre1 in Trichoderma reesei Rut-C30 turns it into an activator Rassinger, Alice Gacek-Matthews, Agnieszka Strauss, Joseph Mach, Robert L. Mach-Aigner, Astrid R. Fungal Biol Biotechnol Research BACKGROUND: The filamentous fungus Trichoderma reesei (T. reesei) is a natural producer of cellulolytic and xylanolytic enzymes and is therefore industrially used. Many industries require high amounts of enzymes, in particular cellulases. Strain improvement strategies by random mutagenesis yielded the industrial ancestor strain Rut-C30. A key property of Rut-C30 is the partial release from carbon catabolite repression caused by a truncation of the repressor Cre1 (Cre1-96). In the T. reesei wild-type strain a full cre1 deletion leads to pleiotropic effects and strong growth impairment, while the truncated cre1-96 enhances cellulolytic activity without the effect of growth deficiencies. However, it is still unclear which function Cre1-96 has in Rut-C30. RESULTS: In this study, we deleted and constitutively expressed cre1-96 in Rut-C30. We found that the presence of Cre1-96 in Rut-C30 is crucial for its cellulolytic and xylanolytic performance under inducing conditions. In the case of the constitutively expressed Cre1-96, the cellulase activity could further be improved approximately twofold. The deletion of cre1-96 led to growth deficiencies and morphological abnormalities. An in silico domain prediction revealed that Cre1-96 has all necessary properties that a classic transactivator needs. Consequently, we investigated the cellular localization of Cre1-96 by fluorescence microscopy using an eYFP-tag. Cre1-96 is localized in the fungal nuclei under both, inducing and repressing conditions. Furthermore, chromatin immunoprecipitation revealed an enrichment of Cre1-96 in the upstream regulatory region of the main transactivator of cellulases and xylanases, Xyr1. Interestingly, transcript levels of cre1-96 show the same patterns as the ones of xyr1 under inducing conditions. CONCLUSIONS: The findings suggest that the truncation turns Cre1 into an activating regulator, which primarily exerts its role by approaching the upstream regulatory region of xyr1. The conversion of repressor proteins to potential activators in other biotechnologically used filamentous fungi can be applied to increase their enzyme production capacities. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1186/s40694-018-0059-0) contains supplementary material, which is available to authorized users. BioMed Central 2018-08-20 /pmc/articles/PMC6100732/ /pubmed/30151221 http://dx.doi.org/10.1186/s40694-018-0059-0 Text en © The Author(s) 2018 Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. |
spellingShingle | Research Rassinger, Alice Gacek-Matthews, Agnieszka Strauss, Joseph Mach, Robert L. Mach-Aigner, Astrid R. Truncation of the transcriptional repressor protein Cre1 in Trichoderma reesei Rut-C30 turns it into an activator |
title | Truncation of the transcriptional repressor protein Cre1 in Trichoderma reesei Rut-C30 turns it into an activator |
title_full | Truncation of the transcriptional repressor protein Cre1 in Trichoderma reesei Rut-C30 turns it into an activator |
title_fullStr | Truncation of the transcriptional repressor protein Cre1 in Trichoderma reesei Rut-C30 turns it into an activator |
title_full_unstemmed | Truncation of the transcriptional repressor protein Cre1 in Trichoderma reesei Rut-C30 turns it into an activator |
title_short | Truncation of the transcriptional repressor protein Cre1 in Trichoderma reesei Rut-C30 turns it into an activator |
title_sort | truncation of the transcriptional repressor protein cre1 in trichoderma reesei rut-c30 turns it into an activator |
topic | Research |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6100732/ https://www.ncbi.nlm.nih.gov/pubmed/30151221 http://dx.doi.org/10.1186/s40694-018-0059-0 |
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