Re-direction of carbon flux to key precursor malonyl-CoA via artificial small RNAs in photosynthetic sp. PCC 6803.

BACKGROUND

Photosynthetic cyanobacteria have attracted a significant attention as promising chassis to produce renewable fuels and chemicals due to their capability to utilizing solar energy and CO. Notably, the enhancing supply of key precursors like malonyl-CoA would benefit the production of many bio-compounds. Nevertheless, the lacking of genetic tools in cyanobacteria, especially the knockdown strategies for essential pathways, has seriously restricted the attempts to re-direct carbon flux from the central carbohydrate metabolism to the synthesis of bioproducts.

RESULTS

Aiming at developing new genetic tools, two small RNA regulatory tools are reported for the model cyanobacterium sp. PCC6803, based on paired termini RNAs as well as the exogenous Hfq chaperone and MicC scaffold (Hfq-MicC) previously developed in . Both regulatory tools functioned well in regulating exogenous reporter gene and endogenous gene in sp. PCC6803, achieving a downregulation of gene expression up to 90% compared with wildtype. In addition, the Hfq-MicC tool was developed to simultaneously regulate multiple genes related to essential fatty acids biosynthesis, which led to decreased fatty acids content by 11%. Furthermore, aiming to re-direct the carbon flux, the Hfq-MicC tool was utilized to interfere the competing pathway of malonyl-CoA, achieving an increased intracellular malonyl-CoA abundance up to 41% (~ 698.3 pg/mL/OD) compared to the wildtype. Finally, the Hfq-MicC system was further modified into an inducible system based on the theophylline-inducible riboswitch.

CONCLUSIONS

In this study, two small RNA regulatory tools for manipulating essential metabolic pathways and re-directing carbon flux are reported for sp. PCC6803. The work introduces efficient and valuable metabolic regulatory strategies for photosynthetic cyanobacteria.