Real-Time Assessment of Intracellular Metabolites in Single Cells through RNA-Based Sensors
2023
Alvaro Darío Ortega
Quantification of the concentration of particular cellular metabolites reports on the actual utilization of metabolic pathways in physiological and pathological conditions. Metabolite concentration also constitutes the readout for screening cell factories in metabolic engineering. However, there are no direct approaches that allow for real-time assessment of the levels of intracellular metabolites in single cells. In recent years, the modular architecture of natural bacterial RNA riboswitches has inspired the design of genetically encoded synthetic RNA devices that convert the intracellular concentration of a metabolite into a quantitative fluorescent signal. These so-called RNA-based sensors are composed of a metabolite-binding RNA aptamer as the sensor domain, connected through an actuator segment to a signal-generating reporter domain. However, at present, the variety of available RNA-based sensors for intracellular metabolites is still very limited. Here, we go through natural mechanisms for metabolite sensing and regulation in cells across all kingdoms, focusing on those mediated by riboswitches. We review the design principles underlying currently developed RNA-based sensors and discuss the challenges that hindered the development of novel sensors and recent strategies to address them. We finish by introducing the current and potential applicability of synthetic RNA-based sensors for intracellular metabolites.
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