Discovery and characterization of Synechocystis sp. PCC 6803 light-entrained promoters in diurnal light:dark cycles

Cyanobacteria are photosynthetic bacteria employed for production of valuable chemicals using sunlight and atmospheric carbon dioxide as substrates. Industrial production in outdoor facilities exposes cyanobacteria to daily light:dark (LD) cycles of sunlight availability. Strain engineers need genetic tools suited for diurnal LD cycles to maximize production in these conditions, but tools for engineering in diurnal LD cycles are extremely limited. Here, we discover native Synechocystis sp. PCC 6803 promoters which provide light-entrained expression in 12-hour light:12-hour dark (12:12) LD cycles. Promoters were characterized using bacterial luciferase bioluminescent promoter probes and RT-qPCR. PhliC, Prbp1, Pslr0006, and PsigA provide light-entrained expression in 12:12 LD cycles when expressed from the slr0168 chromosomal neutral site. None of the promoters provided free-running oscillations in continuous light (CL) following diurnal LD entrainment, but growth in CL resulted in constant mid-level bioluminescence. Transcripts from Prbp1, Pslr0006, and PsigA increased significantly following the onset of light, whereas transcripts from PhliC exhibited no oscillations in 12:12 LD cycles. Furthermore, PhliC bioluminescence induction at the onset of light increased at higher light intensities. Overall, our work provides additional genetic engineering tools for cyanobacterial strains for chemical production in diurnal LD cycles.