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Bidirectional histone-gene promoters in Aspergillus: characterization and application for multi-gene expression

BACKGROUND: Filamentous fungi are important producers of enzymes and bioactive secondary metabolites and are exploited for industrial purposes. Expression and characterization of biosynthetic pathways requires stable expression of multiple genes in the production host. Fungal promoters are indispens...

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Autores principales: Rendsvig, Jakob K. H., Workman, Christopher T., Hoof, Jakob B.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: BioMed Central 2019
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6900853/
https://www.ncbi.nlm.nih.gov/pubmed/31867115
http://dx.doi.org/10.1186/s40694-019-0088-3
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author Rendsvig, Jakob K. H.
Workman, Christopher T.
Hoof, Jakob B.
author_facet Rendsvig, Jakob K. H.
Workman, Christopher T.
Hoof, Jakob B.
author_sort Rendsvig, Jakob K. H.
collection PubMed
description BACKGROUND: Filamentous fungi are important producers of enzymes and bioactive secondary metabolites and are exploited for industrial purposes. Expression and characterization of biosynthetic pathways requires stable expression of multiple genes in the production host. Fungal promoters are indispensable for the accomplishment of this task, and libraries of promoters that show functionality across diverse fungal species facilitate synthetic biology approaches, pathway expression, and cell-factory construction. RESULTS: In this study, we characterized the intergenic region between the genes encoding histones H4.1 and H3, from five phylogenetically diverse species of Aspergillus, as bidirectional promoters (Ph4h3). By expression of the genes encoding fluorescent proteins mRFP1 and mCitrine, we show at the translational and transcriptional level that this region from diverse species is applicable as strong and constitutive bidirectional promoters in Aspergillus nidulans. Bioinformatic analysis showed that the divergent gene orientation of h4.1 and h3 appears maintained among fungi, and that the Ph4h3 display conserved DNA motifs among the investigated 85 Aspergilli. Two of the heterologous Ph4h3s were utilized for single-locus expression of four genes from the putative malformin producing pathway from Aspergillus brasiliensis in A. nidulans. Strikingly, heterologous expression of mlfA encoding the non-ribosomal peptide synthetase is sufficient for biosynthesis of malformins in A. nidulans, which indicates an iterative use of one adenylation domain in the enzyme. However, this resulted in highly stressed colonies, which was reverted to a healthy phenotype by co-expressing the residual four genes from the putative biosynthetic gene cluster. CONCLUSIONS: Our study has documented that Ph4h3 is a strong constitutive bidirectional promoter and a valuable new addition to the genetic toolbox of at least the genus Aspergillus.
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spelling pubmed-69008532019-12-20 Bidirectional histone-gene promoters in Aspergillus: characterization and application for multi-gene expression Rendsvig, Jakob K. H. Workman, Christopher T. Hoof, Jakob B. Fungal Biol Biotechnol Research BACKGROUND: Filamentous fungi are important producers of enzymes and bioactive secondary metabolites and are exploited for industrial purposes. Expression and characterization of biosynthetic pathways requires stable expression of multiple genes in the production host. Fungal promoters are indispensable for the accomplishment of this task, and libraries of promoters that show functionality across diverse fungal species facilitate synthetic biology approaches, pathway expression, and cell-factory construction. RESULTS: In this study, we characterized the intergenic region between the genes encoding histones H4.1 and H3, from five phylogenetically diverse species of Aspergillus, as bidirectional promoters (Ph4h3). By expression of the genes encoding fluorescent proteins mRFP1 and mCitrine, we show at the translational and transcriptional level that this region from diverse species is applicable as strong and constitutive bidirectional promoters in Aspergillus nidulans. Bioinformatic analysis showed that the divergent gene orientation of h4.1 and h3 appears maintained among fungi, and that the Ph4h3 display conserved DNA motifs among the investigated 85 Aspergilli. Two of the heterologous Ph4h3s were utilized for single-locus expression of four genes from the putative malformin producing pathway from Aspergillus brasiliensis in A. nidulans. Strikingly, heterologous expression of mlfA encoding the non-ribosomal peptide synthetase is sufficient for biosynthesis of malformins in A. nidulans, which indicates an iterative use of one adenylation domain in the enzyme. However, this resulted in highly stressed colonies, which was reverted to a healthy phenotype by co-expressing the residual four genes from the putative biosynthetic gene cluster. CONCLUSIONS: Our study has documented that Ph4h3 is a strong constitutive bidirectional promoter and a valuable new addition to the genetic toolbox of at least the genus Aspergillus. BioMed Central 2019-12-09 /pmc/articles/PMC6900853/ /pubmed/31867115 http://dx.doi.org/10.1186/s40694-019-0088-3 Text en © The Author(s) 2019 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
Rendsvig, Jakob K. H.
Workman, Christopher T.
Hoof, Jakob B.
Bidirectional histone-gene promoters in Aspergillus: characterization and application for multi-gene expression
title Bidirectional histone-gene promoters in Aspergillus: characterization and application for multi-gene expression
title_full Bidirectional histone-gene promoters in Aspergillus: characterization and application for multi-gene expression
title_fullStr Bidirectional histone-gene promoters in Aspergillus: characterization and application for multi-gene expression
title_full_unstemmed Bidirectional histone-gene promoters in Aspergillus: characterization and application for multi-gene expression
title_short Bidirectional histone-gene promoters in Aspergillus: characterization and application for multi-gene expression
title_sort bidirectional histone-gene promoters in aspergillus: characterization and application for multi-gene expression
topic Research
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6900853/
https://www.ncbi.nlm.nih.gov/pubmed/31867115
http://dx.doi.org/10.1186/s40694-019-0088-3
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