Regulation Of Ethanol Tolerance By A Transcriptional Regulator Sll0794 In Synechocystis Sp. PCC 6803

Biofuel ethanol can be produced directly from CO2 in a model photosynthetic cyanobacterium Synechocystis sp. PCC 6803. The progress in synthetic biology has led to autotrophic production of ethanol in various cyanobacterial hosts. However, cyanobacteria have very low tolerance to ethanol, which restricts the economic viability of ethanol production from these renewable host systems. Therefore, it is of great significance to explore regulatory mechanisms of ethanol tolerance so that engineering more robust ethanol-producing cyanobacterial hosts can be possible.Our previous proteomic and transcriptomic analyses found that several regulatory proteins were up regulated in Synechocystis sp. PCC 6803 after ethanol treatment. In this study, through tolerance analysis of the gene disruption mutants of three up-regulated regulatory genes, we uncovered that one transcriptional regulator, Sll0794, was related directly to ethanol tolerance in Synechocystis. Using a quantitative iTRAQ-LC-MS/MS proteomics approach, we further determined the possible regulatory network of Sll0794. The proteomic analysis showed that a total of 54 and 87 unique proteins were down- and up-regulated in the ?sll0794 mutant grown under ethanol stress, respectively. In addition, electrophoretic mobility shift assays(EMSAs) demonstrated that the Sll0794 transcriptional regulator was able to bind directly to the upstream regions of sll1514, slr1512 and slr1838, which encode a 16.6 kDa small heat shock protein, a putative sodium-dependent bicarbonate transporter and a carbon dioxide concentrating mechanism protein CcmK, respectively.The study provided a proteomic description of the putative ethanol-tolerance network regulated by the sll0794 gene, and revealed new insights on the ethanol-tolerance regulatory mechanism in Synechocystis. As the first regulatory protein discovered related to ethanol tolerance, the gene can serve as a valuable target for transcriptional engineering in order to improve ethanol tolerance in Synechocystis.