Genetic analysis of the microcystin biosynthesis gene cluster in Microcystis strains from four bodies of eutrophic water in Japan

• Published in: Journal of general and applied microbiology Vol. 55; no. 2; pp. 111 - 123
• Main Authors: Noguchi, Takahiko, Shinohara, Azusa, Nishizawa, Akito, Asayama, Munehiko, Nakano, Tomoyo, Hasegawa, Masateru, Harada, Ken-ichi, Nishizawa, Tomoyasu, Shirai, Makoto
• Format: Journal Article
• Language: English
• Published: Tokyo Applied Microbiology, Molecular and Cellular Biosciences Research Foundation 2009; Microbiology Research Foundation; Japan Science and Technology Agency
• Subjects: 16S-23S rRNA gene ITS sequence; Bacterial Proteins - genetics; Base Sequence; Biological and medical sciences; cyanobacteria; DNA, Ribosomal Spacer - genetics; Fundamental and applied biological sciences. Psychology; Gene Expression Regulation, Bacterial; Gene Order; Genes, Bacterial - genetics; Japan; McyI dehydrogenase activity; Microbiology; microcystin biosynthesis gene; Microcystins - genetics; Microcystis; Microcystis - classification; Microcystis - genetics; Microcystis - metabolism; Molecular Sequence Data; Multigene Family - genetics; Peptide Synthases - biosynthesis; Peptide Synthases - genetics; Phylogeny; Water Microbiology; Asia; Japan; Water; Gene cluster; Nucleotide sequence; Enzyme; Biosynthesis; Dehydrogenase; Microcystis; Gene; Ribosomal RNA; Cyanobacteria; 16S-RNA; Bacteria; Genetics; Mcyl dehydrogenase activity; Oxidoreductases; microcystin biosynthesis gene; 16S-23S rRNA gene ITS sequence
• ISSN: 0022-1260; 1349-8037
• DOI: 10.2323/jgam.55.111

The highly conserved organization of microcystin biosynthesis (mcy) gene clusters, which includes nonribosomal peptide synthetase (NRPS) genes, polyketide synthase (PKS) genes, and fused NRPS-PKS genes, has been characterized in the genus Microcystis. In this study, a total of 135 cyanobacterial strains from four different geographical locations in Japan were isolated. Fourteen mcy-possessing (mcy+) strains were identified according to PCR amplification between two genes from domestic mcy+ strains and the mcy gene’s organization was classified into five types. Phylogenetic relationships of the 16S-23S internal transcribed spacer region indicated that the five types of mcy gene cluster structure classified into two groups of the genus Microcystis. HPLC of the isolated mcy+ strain containing a partial deletion of mcyI (ΔmcyI) revealed that microcystin production disappeared. A transcriptional analysis of the Δ mcyI-strain and an assay of recombinant McyI dehydrogenase activity showed that McyI is responsible for microcystin biosynthesis. Based on patterns of the PCR amplicons and analyses of nucleotide sequences in the mcy gene cluster of Microcystis, we confirmed the presence of inserts at three specific loci, between mcyA and mcyD, and downstream of mcyC and mcyJ. Our study is the first investigation of the mcy gene cluster structure in the genus Microcystis from environmental samples.