Investigation of singlet-oxygen-responsive genes in the cyanobacterium Synechocystis PCC 6803.

Singlet oxygen (1O2) is an important reactive oxygen species whose formation by the type-II, light-dependent, photodynamic reaction is inevitable during photosynthetic processes. In the last decades, the recognition that 1O2 is not only a damaging agent, but can also affect gene expression and participates in signal transduction pathways has received increasing attention. However, contrary to several other taxa, 1O2-responsive genes have not been identified in the important cyanobacterial model organism Synechocystis PCC 6803. By using global transcript analysis we have identified a large set of Synechocystis genes, whose transcript levels were either enhanced or repressed in the presence of 1O2. Characteristic 1O2 responses were observed in several light-inducible genes of Synechocystis, especially in the hli (or scp) family encoding HLIP/SCP proteins involved in photoprotection. Other important 1O2-induced genes include components of the Photosystem II repair machinery (psbA2 and ftsH2, ftsH3), iron homeostasis genes isiA and idiA, the group 2 sigma factor sigD, some components of the transcriptomes induced by salt-, hyperosmotic and cold-stress, as well as several genes of unknown function. The most pronounced 1O2-induced upregulation was observed for the hliB and the co-transcribed lilA genes, whose deletion induced enhanced sensitivity against 1O2-mediated light damage. A bioreporter Synechocystis strain was created by fusing the hliB promoter to the bacterial luciferase (lux), which showed its utility for continuous monitoring of 1O2 concentrations inside the cell.

Impact of Carbon Fixation, Distribution and Storage on the Production of Farnesene and Limonene in Synechocystis PCC 6803 and Synechococcus PCC 7002.

Terpenes are high-value chemicals which can be produced by engineered cyanobacteria from sustainable resources, solar energy, water and CO2. We previously reported that the euryhaline unicellular cyanobacteria Synechocystis sp. PCC 6803 (S.6803) and Synechococcus sp. PCC 7002 (S.7002) produce farnesene and limonene, respectively, more efficiently than other terpenes. In the present study, we attempted to enhance farnesene production in S.6803 and limonene production in S.7002. Practically, we tested the influence of key cyanobacterial enzymes acting in carbon fixation (RubisCO, PRK, CcmK3 and CcmK4), utilization (CrtE, CrtR and CruF) and storage (PhaA and PhaB) on terpene production in S.6803, and we compared some of the findings with the data obtained in S.7002. We report that the overproduction of RubisCO from S.7002 and PRK from Cyanothece sp. PCC 7425 increased farnesene production in S.6803, but not limonene production in S.7002. The overexpression of the crtE genes (synthesis of terpene precursors) from S.6803 or S.7002 did not increase farnesene production in S.6803. In contrast, the overexpression of the crtE gene from S.6803, but not S.7002, increased farnesene production in S.7002, emphasizing the physiological difference between these two model cyanobacteria. Furthermore, the deletion of the crtR and cruF genes (carotenoid synthesis) and phaAB genes (carbon storage) did not increase the production of farnesene in S.6803. Finally, as a containment strategy of genetically modified strains of S.6803, we report that the deletion of the ccmK3K4 genes (carboxysome for CO2 fixation) did not affect the production of limonene, but decreased the production of farnesene in S.6803.