CRISPR-Cas9-Based Discovery of the Verrucosidin Biosynthesis Gene Cluster in Penicillium polonicum
Penicillium polonicum, commonly found on food matrices, is a mycotoxigenic species able to produce a neurotoxin called verrucosidin. This methylated α-pyrone polyketide inhibits oxidative phosphorylation in mitochondria and thereby causes neurological diseases. Despite the importance of verrucosidin...
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| Format: | Article |
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Frontiers Media S.A.
2021-05-01
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| Series: | Frontiers in Microbiology |
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| Online Access: | https://www.frontiersin.org/articles/10.3389/fmicb.2021.660871/full |
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| author | Silvia Valente Silvia Valente Edoardo Piombo Edoardo Piombo Volker Schroeckh Giovanna Roberta Meloni Giovanna Roberta Meloni Thorsten Heinekamp Axel A. Brakhage Axel A. Brakhage Davide Spadaro Davide Spadaro |
| author_facet | Silvia Valente Silvia Valente Edoardo Piombo Edoardo Piombo Volker Schroeckh Giovanna Roberta Meloni Giovanna Roberta Meloni Thorsten Heinekamp Axel A. Brakhage Axel A. Brakhage Davide Spadaro Davide Spadaro |
| author_sort | Silvia Valente |
| collection | DOAJ |
| description | Penicillium polonicum, commonly found on food matrices, is a mycotoxigenic species able to produce a neurotoxin called verrucosidin. This methylated α-pyrone polyketide inhibits oxidative phosphorylation in mitochondria and thereby causes neurological diseases. Despite the importance of verrucosidin as a toxin, its biosynthetic genes have not been characterized yet. By similarity analysis with the polyketide synthase (PKS) genes for the α-pyrones aurovertin (AurA) and citreoviridin (CtvA), 16 PKS genes for putative α-pyrones were identified in the P. polonicum genome. A single PKS gene, verA, was found to be transcribed under verrucosidin-producing growth conditions. The annotated functions of the genes neighboring verA correspond to those required for verrucosidin biosynthesis. To prove the involvement of verA in verrucosidin biosynthesis, the clustered regularly interspaced short palindrome repeats (CRISPR) technology was applied to P. polonicum. In vitro reconstituted CRISPR-Cas9 was used to induce targeted gene deletions in P. polonicum. This approach allowed identifying and characterizing the verrucosidin biosynthetic gene cluster. VerA deletion mutants were no longer able to produce verrucosidin, whereas they were displaying morphological characteristics comparable with the wild-type strain. The available CRISPR-Cas9 technology allows characterizing the biosynthetic potential of P. polonicum as a valuable source of novel compounds. |
| first_indexed | 2024-12-16T23:08:57Z |
| format | Article |
| id | doaj.art-39eb35cd08be469ebccc44743e9c89f0 |
| institution | Directory Open Access Journal |
| issn | 1664-302X |
| language | English |
| last_indexed | 2024-12-16T23:08:57Z |
| publishDate | 2021-05-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Frontiers in Microbiology |
| spelling | doaj.art-39eb35cd08be469ebccc44743e9c89f02022-12-21T22:12:28ZengFrontiers Media S.A.Frontiers in Microbiology1664-302X2021-05-011210.3389/fmicb.2021.660871660871CRISPR-Cas9-Based Discovery of the Verrucosidin Biosynthesis Gene Cluster in Penicillium polonicumSilvia Valente0Silvia Valente1Edoardo Piombo2Edoardo Piombo3Volker Schroeckh4Giovanna Roberta Meloni5Giovanna Roberta Meloni6Thorsten Heinekamp7Axel A. Brakhage8Axel A. Brakhage9Davide Spadaro10Davide Spadaro11AGROINNOVA – Centre of Competence for the Innovation in the Agro-Environmental Sector, Grugliasco, ItalyDepartment of Agricultural, Forest and Food Sciences, Università degli Studi di Torino, Grugliasco, ItalyAGROINNOVA – Centre of Competence for the Innovation in the Agro-Environmental Sector, Grugliasco, ItalyDepartment of Agricultural, Forest and Food Sciences, Università degli Studi di Torino, Grugliasco, ItalyDepartment of Molecular and Applied Microbiology, Leibniz Institute for Natural Product Research and Infection Biology – Hans Knöll Institute, Jena, GermanyAGROINNOVA – Centre of Competence for the Innovation in the Agro-Environmental Sector, Grugliasco, ItalyDepartment of Agricultural, Forest and Food Sciences, Università degli Studi di Torino, Grugliasco, ItalyDepartment of Molecular and Applied Microbiology, Leibniz Institute for Natural Product Research and Infection Biology – Hans Knöll Institute, Jena, GermanyDepartment of Molecular and Applied Microbiology, Leibniz Institute for Natural Product Research and Infection Biology – Hans Knöll Institute, Jena, GermanyDepartment of Microbiology and Molecular Biology, Institute for Microbiology, Friedrich Schiller University, Jena, GermanyAGROINNOVA – Centre of Competence for the Innovation in the Agro-Environmental Sector, Grugliasco, ItalyDepartment of Agricultural, Forest and Food Sciences, Università degli Studi di Torino, Grugliasco, ItalyPenicillium polonicum, commonly found on food matrices, is a mycotoxigenic species able to produce a neurotoxin called verrucosidin. This methylated α-pyrone polyketide inhibits oxidative phosphorylation in mitochondria and thereby causes neurological diseases. Despite the importance of verrucosidin as a toxin, its biosynthetic genes have not been characterized yet. By similarity analysis with the polyketide synthase (PKS) genes for the α-pyrones aurovertin (AurA) and citreoviridin (CtvA), 16 PKS genes for putative α-pyrones were identified in the P. polonicum genome. A single PKS gene, verA, was found to be transcribed under verrucosidin-producing growth conditions. The annotated functions of the genes neighboring verA correspond to those required for verrucosidin biosynthesis. To prove the involvement of verA in verrucosidin biosynthesis, the clustered regularly interspaced short palindrome repeats (CRISPR) technology was applied to P. polonicum. In vitro reconstituted CRISPR-Cas9 was used to induce targeted gene deletions in P. polonicum. This approach allowed identifying and characterizing the verrucosidin biosynthetic gene cluster. VerA deletion mutants were no longer able to produce verrucosidin, whereas they were displaying morphological characteristics comparable with the wild-type strain. The available CRISPR-Cas9 technology allows characterizing the biosynthetic potential of P. polonicum as a valuable source of novel compounds.https://www.frontiersin.org/articles/10.3389/fmicb.2021.660871/fullsecondary metabolitesPenicilliummycotoxinsCRISPR-Casalpha-pyrone polyketides |
| spellingShingle | Silvia Valente Silvia Valente Edoardo Piombo Edoardo Piombo Volker Schroeckh Giovanna Roberta Meloni Giovanna Roberta Meloni Thorsten Heinekamp Axel A. Brakhage Axel A. Brakhage Davide Spadaro Davide Spadaro CRISPR-Cas9-Based Discovery of the Verrucosidin Biosynthesis Gene Cluster in Penicillium polonicum Frontiers in Microbiology secondary metabolites Penicillium mycotoxins CRISPR-Cas alpha-pyrone polyketides |
| title | CRISPR-Cas9-Based Discovery of the Verrucosidin Biosynthesis Gene Cluster in Penicillium polonicum |
| title_full | CRISPR-Cas9-Based Discovery of the Verrucosidin Biosynthesis Gene Cluster in Penicillium polonicum |
| title_fullStr | CRISPR-Cas9-Based Discovery of the Verrucosidin Biosynthesis Gene Cluster in Penicillium polonicum |
| title_full_unstemmed | CRISPR-Cas9-Based Discovery of the Verrucosidin Biosynthesis Gene Cluster in Penicillium polonicum |
| title_short | CRISPR-Cas9-Based Discovery of the Verrucosidin Biosynthesis Gene Cluster in Penicillium polonicum |
| title_sort | crispr cas9 based discovery of the verrucosidin biosynthesis gene cluster in penicillium polonicum |
| topic | secondary metabolites Penicillium mycotoxins CRISPR-Cas alpha-pyrone polyketides |
| url | https://www.frontiersin.org/articles/10.3389/fmicb.2021.660871/full |
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