(620k) Engineered Riboswitch: An Effective Way to Control Gene Expression in Cyanobacteria

Authors: 
Saha, R., Washington University
Berla, B. M., Washington University in St. Louis
Pakrasi, H. B., Washington University

Cyanobacteria are photosynthetic microbes that can derive energy from sunlight and carbon from atmospheric carbon dioxide, as well as other sources. In the past two decades, there were numerous efforts geared towards developing cyanobacteria as bioproduction platforms. The majority of these organisms are amenable to genetic manipulation, thus facilitating synthetic biology/metabolic engineering applications. The unique demands of living on light as a primary nutrient make the use of many established gene expression control systems difficult or impossible to apply. Among other approaches, researchers have used endogenous promoters such as PpsbA2 and PcpcB, and synthetic riboswitches with limited/mixed success. Hence, developing an effective genetic toolkit to regulate gene expressions in cyanobacteria is of utmost importance. In this work, we combined a previously developed theophylline-responsive riboswitch with the T7 control system to drive expression of a yellow fluorescence protein (YFP) reporter in Synechocystis sp. PCC 6803. We then demonstrated superiority of this control system over available synthetic riboswitches and isopropyl-β-D-thiogalactopyranoside (IPTG) induced lacIq-Ptrc promoter system. Thus, our work shows the utility of an engineered riboswitch system that can exploit the differences in light/dark life cycle and expedite biotechnological applications in cyanobacteria.