Castillo-Hair, Sebastian M.Baerman, Elliot A.Fujita, MasayaIgoshin, Oleg A.Tabor, Jeffrey J.2019-11-182019-11-182019Castillo-Hair, Sebastian M., Baerman, Elliot A., Fujita, Masaya, et al.. "Optogenetic control of Bacillus subtilis gene expression." <i>Nature Communications,</i> 10, (2019) Springer Nature: https://doi.org/10.1038/s41467-019-10906-6.https://hdl.handle.net/1911/107700The Gram-positive bacterium Bacillus subtilis exhibits complex spatial and temporal gene expression signals. Although optogenetic tools are ideal for studying such processes, none has been engineered for this organism. Here, we port a cyanobacterial light sensor pathway comprising the green/red photoreversible two-component system CcaSR, two metabolic enzymes for production of the chromophore phycocyanobilin (PCB), and an output promoter to control transcription of a gene of interest into B. subtilis. Following an initial non-functional design, we optimize expression of pathway genes, enhance PCB production via a translational fusion of the biosynthetic enzymes, engineer a strong chimeric output promoter, and increase dynamic range with a miniaturized photosensor kinase. Our final design exhibits over 70-fold activation and rapid response dynamics, making it well-suited to studying a wide range of gene regulatory processes. In addition, the synthetic biology methods we develop to port this pathway should make B. subtilis easier to engineer in the future.engThis article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder.Journal articles41467-019-10906-6https://doi.org/10.1038/s41467-019-10906-6