Inactivation of an ABC transporter gene, mcyH, results in loss of microcystin production in the cyanobacterium Microcystis aeruginosa PCC 7806.
The cyanobacterium Microcystis aeruginosa is widely known for its production of the potent hepatotoxin microcystin. Microcystin is synthesized nonribosomally by the thiotemplate function of a large, modular enzyme complex encoded within the 55-kb microcystin synthetase (mcy) gene cluster. Also encoded within the mcy gene cluster is a putative ATP binding cassette (ABC) transporter, McyH. This study details the bioinformatic and mutational analyses of McyH and offers functional predictions for the hypothetical protein. The transporter is putatively comprised of two homodimers, each with an N-terminal hydrophobic domain and a C-terminal ATPase. Phylogenetically, McyH was found to cluster with members of the ABC-A1 subgroup of ABC ATPases, suggesting an export function for the protein. Two mcyH null mutant (DeltamcyH) strains were constructed by partial deletion of the mcyH gene. Microcystin production was completely absent in these strains. While the mcyH deletion had no apparent effect on the transcription of other mcy genes, the complete microcystin biosynthesis enzyme complex could not be detected in DeltamcyH mutant strains. Finally, expression levels of McyH in the wild type and in DeltamcyA, DeltamcyB, and DeltamcyH mutants were investigated by using immunoblotting with an anti-McyH antibody. Expression of McyH was found to be reduced in DeltamcyA and DeltamcyB mutants and completely absent in the DeltamcyH mutant. By virtue of its association with the mcy gene cluster and the bioinformatic and experimental data presented in this study, we predict that McyH functions as a microcystin exporter and is, in addition, intimately associated with the microcystin biosynthesis pathway.
Pearson
Leanne A
LA
School of Biotechnology and Biomolecular Sciences, The University of New South Wales, Sydney, Australia.
Hisbergues
Michael
M
Börner
Thomas
T
Dittmann
Elke
E
Neilan
Brett A
BA
D018528:ATP-Binding Cassette Transporters / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D001427:Bacterial Toxins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D017353:Gene Deletion; D052998:Microcystins; D046931:Microcystis / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D008969:Molecular Sequence Data; D005810:Multigene Family; D010456:Peptides, Cyclic / Q000096:biosynthesis*; D010802:Phylogeny; D017422:Sequence Analysis, DNA
D018528:ATP-Binding Cassette Transporters; D001426:Bacterial Proteins; D001427:Bacterial Toxins; C496657:McyH protein, Microcystis aeruginosa; D052998:Microcystins; D010456:Peptides, Cyclic; C078588:microcystin