Insertional Mutagenesis and Cloning of the Genes Required for Biosynthesis of the Host-Specific AK-Toxin in the Japanese Pear Pathotype of Alternaria alternata
The Japanese pear pathotype of Alternaria alternata causes black spot of Japanese pear by producing a host-specific toxin known as AK-toxin. Restriction enzyme-mediated integration (REMI) mutagenesis was used to tag genes required for toxin biosynthesis. Protoplasts of a wild-type strain were treate...
| Main Authors: | , , , |
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| Format: | Article |
| Language: | English |
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The American Phytopathological Society
1999-08-01
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| Series: | Molecular Plant-Microbe Interactions |
| Online Access: | https://apsjournals.apsnet.org/doi/10.1094/MPMI.1999.12.8.691 |
| _version_ | 1828264637170188288 |
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| author | Aiko Tanaka Hiroshi Shiotani Mikihiro Yamamoto Takashi Tsuge |
| author_facet | Aiko Tanaka Hiroshi Shiotani Mikihiro Yamamoto Takashi Tsuge |
| author_sort | Aiko Tanaka |
| collection | DOAJ |
| description | The Japanese pear pathotype of Alternaria alternata causes black spot of Japanese pear by producing a host-specific toxin known as AK-toxin. Restriction enzyme-mediated integration (REMI) mutagenesis was used to tag genes required for toxin biosynthesis. Protoplasts of a wild-type strain were treated with a linearized plasmid along with the restriction enzyme used to linearize the plasmid. Of 984 REMI transformants recovered, three produced no detectable AK-toxin and lost pathogenicity on pear leaves. Genomic DNA flanking the integrated plasmid was recovered from one of the mutants. With the recovered DNA used as a probe, a cosmid clone of the wild-type strain was isolated. Structural and functional analyses of an 8.0-kb region corresponding to the tagged site indicated the presence of two genes. One, designated AKT1, encodes a member of the class of carboxyl-activating enzymes. The other, AKT2, encodes a protein of unknown function. The essential roles of these two genes in both AK-toxin production and pathogenicity were confirmed by transformation-mediated gene disruption experiments. DNA gel blot analysis detected AKT1 and AKT2 homologues not only in the Japanese pear pathotype strains but also in strains from the tangerine and strawberry pathotypes. The host-specific toxins of these two pathotypes are similar in structure to AK-toxin. Homologues were not detected in other pathotypes or in non-pathogenic strains of A. alternata, suggesting acquisition of AKT1 and AKT2 by horizontal transfer. |
| first_indexed | 2024-04-13T04:27:49Z |
| format | Article |
| id | doaj.art-a3a2eb169b37423bb028e52e7cb20016 |
| institution | Directory Open Access Journal |
| issn | 0894-0282 1943-7706 |
| language | English |
| last_indexed | 2024-04-13T04:27:49Z |
| publishDate | 1999-08-01 |
| publisher | The American Phytopathological Society |
| record_format | Article |
| series | Molecular Plant-Microbe Interactions |
| spelling | doaj.art-a3a2eb169b37423bb028e52e7cb200162022-12-22T03:02:27ZengThe American Phytopathological SocietyMolecular Plant-Microbe Interactions0894-02821943-77061999-08-0112869170210.1094/MPMI.1999.12.8.691Insertional Mutagenesis and Cloning of the Genes Required for Biosynthesis of the Host-Specific AK-Toxin in the Japanese Pear Pathotype of Alternaria alternataAiko TanakaHiroshi ShiotaniMikihiro YamamotoTakashi TsugeThe Japanese pear pathotype of Alternaria alternata causes black spot of Japanese pear by producing a host-specific toxin known as AK-toxin. Restriction enzyme-mediated integration (REMI) mutagenesis was used to tag genes required for toxin biosynthesis. Protoplasts of a wild-type strain were treated with a linearized plasmid along with the restriction enzyme used to linearize the plasmid. Of 984 REMI transformants recovered, three produced no detectable AK-toxin and lost pathogenicity on pear leaves. Genomic DNA flanking the integrated plasmid was recovered from one of the mutants. With the recovered DNA used as a probe, a cosmid clone of the wild-type strain was isolated. Structural and functional analyses of an 8.0-kb region corresponding to the tagged site indicated the presence of two genes. One, designated AKT1, encodes a member of the class of carboxyl-activating enzymes. The other, AKT2, encodes a protein of unknown function. The essential roles of these two genes in both AK-toxin production and pathogenicity were confirmed by transformation-mediated gene disruption experiments. DNA gel blot analysis detected AKT1 and AKT2 homologues not only in the Japanese pear pathotype strains but also in strains from the tangerine and strawberry pathotypes. The host-specific toxins of these two pathotypes are similar in structure to AK-toxin. Homologues were not detected in other pathotypes or in non-pathogenic strains of A. alternata, suggesting acquisition of AKT1 and AKT2 by horizontal transfer.https://apsjournals.apsnet.org/doi/10.1094/MPMI.1999.12.8.691 |
| spellingShingle | Aiko Tanaka Hiroshi Shiotani Mikihiro Yamamoto Takashi Tsuge Insertional Mutagenesis and Cloning of the Genes Required for Biosynthesis of the Host-Specific AK-Toxin in the Japanese Pear Pathotype of Alternaria alternata Molecular Plant-Microbe Interactions |
| title | Insertional Mutagenesis and Cloning of the Genes Required for Biosynthesis of the Host-Specific AK-Toxin in the Japanese Pear Pathotype of Alternaria alternata |
| title_full | Insertional Mutagenesis and Cloning of the Genes Required for Biosynthesis of the Host-Specific AK-Toxin in the Japanese Pear Pathotype of Alternaria alternata |
| title_fullStr | Insertional Mutagenesis and Cloning of the Genes Required for Biosynthesis of the Host-Specific AK-Toxin in the Japanese Pear Pathotype of Alternaria alternata |
| title_full_unstemmed | Insertional Mutagenesis and Cloning of the Genes Required for Biosynthesis of the Host-Specific AK-Toxin in the Japanese Pear Pathotype of Alternaria alternata |
| title_short | Insertional Mutagenesis and Cloning of the Genes Required for Biosynthesis of the Host-Specific AK-Toxin in the Japanese Pear Pathotype of Alternaria alternata |
| title_sort | insertional mutagenesis and cloning of the genes required for biosynthesis of the host specific ak toxin in the japanese pear pathotype of alternaria alternata |
| url | https://apsjournals.apsnet.org/doi/10.1094/MPMI.1999.12.8.691 |
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