Reconstitution and Expression of mcy Gene Cluster in The Model Cyanobacterium Synechococcus 7942 Reveals a Roll of MC-LR in Cell Division

• Published in: bioRxiv
• Main Authors: Zheng Yanli, Xue Chunling, Chen, Hui, Jia Anqi, Zhao, Liang, Zhang, Junli, Zhang, Lixin, Wang, Qiang
• Format: Paper
• Language: English
• Published: Cold Spring Harbor Cold Spring Harbor Laboratory Press 16.04.2022
• Subjects: Binding sites; Biosynthesis; Cell division; Chronic exposure; Decarboxylation; Filamentation; Hepatocellular carcinoma; Hydrocarbons; Microcystins; Phenylpropanoids; Public health; Recombination; Synechococcus
• DOI: 10.1101/2022.02.17.480855

Cyanobacterial blooms pose a serious threat to public health due to the presence of cyanotoxins. The most common cyanotoxins, microcystins (MCs), can cause acute poisoning at high concentrations and hepatocellular carcinoma following chronic exposure. Among all MC variants, MC-LR produced by Microcystis aeruginosa PCC 7806 is the most common toxic MC. Although the biosynthetic pathway for MC-LR has been proposed, experimental support of this pathway is lacking. In an effort to experimentally validate this pathway, we expressed the 55 kb microcystin biosynthetic gene cluster (mcy cluster) (mcyA-J) and produced MC-LR in the model cyanobacterium Synechococcus 7942. We designed and constructed the strong bidirectional promoter biPpsbA2 between mcyA and mcyD, reassembled the mcy cluster in yeast by transformation-associated recombination (TAR cloning), transformed the gene cluster into the NSII site of Synechococcus 7942, and successfully expressed MC-LR at a level of 0.006-0.018 fg cell-1 day-1. The expression of MC-LR led to abnormal cell division and the filamentation of Synechococcus 7942 cells, further analysis proved a role of MC-LR in functional assembly of the cell division protein FtsZ, by competing its GTP binding site. These results represent the first synthetic biological expression of the mcy cluster and the autotrophic production of MC-LR in a photosynthetic model organism, which lays the foundation for resolving the MC biosynthesis pathway. The suggested role of MC-LR in cell division reveals a mechanism of how blooming cyanobacteria gain a competitive edge over their non-blooming counterparts. Competing Interest Statement The authors have declared no competing interest. Footnotes * 1. The sentence,therefore, phenylpropanoids, the initial substrate of microcystin biosynthesis, may undergo decarboxylation during the first hydrocarbon chain extension reaction, but the specific reaction process is unclear. Furthermore, some intermediate steps in microcystin biosynthesis are also unclear, will be modified to Therefore, phenylpropanoids, the initial substrate of microcystin biosynthesis, may undergo decarboxylation during the first hydrocarbon chain extension reaction, however, the specific reaction process and some intermediate steps in microcystin biosynthesis pathway are Unclear. 2. The homogeneous references have checked carefully and have revised. 3. The funding has been updated.