From Dusk till Dawn: One-plasmid systems for light-regulated gene expression

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Robert Ohlendorf, Andreas Mglich, Keith Moffat

Humboldt-Universitt zu Berlin, Germany

For the quantitative investigation of intracellular signaling and metabolic networks besides precise observa- tional techniques there is a need for perturbative tools featuring high spatiotemporal resolution and minimum invasiveness. Using genetically encoded photoreceptors, optogenetics offers these features. Based on the blue- light-sensitive YF1 histidine kinase (Mglich et al. JMB (2009) 385, 1433-1444) we implemented light-regulated gene expression in Escherichia coli. We have built two systems, pDusk and pDawn, facilitating suppression or activation of gene expression upon blue-light illumination in a dose-dependent manner, with high dynamic range and up to 460-fold induction, respectively. Furthermore pDusk and pDawn are realized on single plasmids and use a flavin-based photoreceptor, therefore offering good portability and independence of external chromophore supply. Both systems constitute valuable tools suitable for basic research as well as for recombinant protein production. Moreover they allow light control of single nodes of signaling networks to elucidate their function within the cellular context.