Abstract
Controlling gene expression by light with fine spatiotemporal resolution not only allows understanding and manipulating fundamental biological processes but also fuels the development of novel therapeutic strategies. In complement to exploiting optogenetic tools, photochemical strategies mostly rely on the incorporation of photo-responsive small molecules into the corresponding biomacromolecular scaffolds. Therefore, generally large synthetic effort is required and the switching of gene expression in both directions within a single system remains a challenge. Here, we report a trans encoded ribo-switch, which consists of an engineered tRNA mimicking structure (TMS), under control of small photo-switchable signalling molecules. The signalling molecules consist of two amino glycoside molecules that are connected via an azobenzene unit. The light responsiveness of our system originates from the photo-switchable noncovalent interactions between the signalling molecule and the TMS switch, leading to the demonstration of photochemically controlled expression of two different genes. We believe that this modular design will provide a powerful platform for controlling the expression of other functional proteins with high spatiotemporal resolution employing light as a stimulus.
Highlights
Synthetic biology aims to develop genetic circuits to reprogram cell behaviour.[1,2] These complex genetic parts that allow perturbing and interpreting biological processes usually function through reacting to exogenous chemical inducers.[3,4] these chemical inducers generally result in a constitutive effect, i.e., in a continuous “ON” state, which limits their application to study inherently dynamic behaviour of cells.[5]
In order to design the photo switchable ligand binding to the tRNA mimicking structure (TMS) switch, we decided to take paromomycin as the parent compound for further modi cations
We have demonstrated a powerful photoresponsive gene expression system that relies on a TMS switch and the corresponding photo-switchable F-dimer ligand
Summary
Synthetic biology aims to develop genetic circuits to reprogram cell behaviour.[1,2] These complex genetic parts that allow perturbing and interpreting biological processes usually function through reacting to exogenous chemical inducers.[3,4] these chemical inducers generally result in a constitutive effect, i.e., in a continuous “ON” state, which limits their application to study inherently dynamic behaviour of cells.[5]. One is the application of optogenetic tools, which generally control protein expression by regulating the interaction between a photo controllable transcription factor and its promoter through light irradiation.[8,9] This approach necessitates the expression of additional photoresponsive proteins which might cause burden on metabolic and cell signaling pathways.[10] The second photochemical approach[11] relies on the installation of light responsive small molecules onto the bio-macromolecular scaffolds including nucleic acids[12,13] and proteins,[14,15,16] thereby providing an extra layer of control over their biological functions by light. This approach generally requires large synthetic efforts for covalently modifying the bio-macromolecules with photo-caging groups[17] or photo-switches.[18,19] the utilization of photocages only permits a single-way gene regulation event due to the deprotective removal of the caging groups under light illumination.[20]
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