Photocaged functional nucleic acids for spatiotemporal imaging in biology
Graphical abstract
Introduction
Biological processes build upon many delicate regulatory events of molecules from nucleic acids and proteins to organic metabolites and metal ions, whose concentrations vary widely with time and location in cells or other biological systems. Consequently, detecting, visualizing, and controlling these molecules in living organisms with high spatiotemporal resolution is very important in our understanding of biological processes. To achieve this goal, photocaged molecules, which are inactive in the absence of light and then activated upon light irradiation, have emerged as perhaps the most effective method because light activation is both kinetically fast and has high spatial resolution, allowing control of both the timing and location of the activation. A primary example is optogenetics in which precise modulation of intracellular signaling in intact cells and multicellular organisms has been achieved at subcellular resolution within seconds of light irradiation [1,2]. While optogenetics has been applied to control the functions of nucleic acids and proteins, it has been challenging to detect and control small organic metabolites and metal ions.
To overcome the limitation in the controlled detection of small molecule metabolites and metal ions, probes based on functional nucleic acids (FNAs) have been developed. FNAs are nucleic acids with either catalytic activities, called catalytic nucleic acids, including ribozymes [3,4] and deoxyribozymes (DNAzymes) [5], or selective binding ability, called riboswitches [6] and DNA aptamers [7,8]. The FNAs can be obtained using a combinatorial process called in vitro selection or Systematic Evolution of Ligands by EXponential enrichment from a large DNA or RNA library of up to 1015 random sequences to bind not only large biomolecules like proteins but also small molecules like metabolites and metal ions [7, 8, 9]. After obtaining FNAs that can bind the targets selectively, signal transducers such as fluorophores can be conjugated to these FNAs to transform them into different in vitro sensors or in vivo imaging agents [10, 11, 12, 13, 14, 15, 16∗,17,∗∗18,19∗∗, 20, 21∗].
While FNA sensors and imaging agents have significantly expanded the number of targets of biological probes, because it takes time to deliver FNAs into cells and other living organisms, the sensing action may occur during the delivery process. To overcome this limitation, FNAs can be protected using photocaged groups. In this way, the function of FNAs is inhibited during the delivery process. Once the FNAs reach the desired location within the living organism, light irradiation can remove the caging group and reactivate the FNAs. More importantly, through control of the timing and location of the light irradiation, spatiotemporal control of the FNA sensors is achieved.
In this review, we summarize photocaging methods and strategies for spatiotemporal control of catalytic nucleic acids and aptamers and how these photocages facilitate regulation of the FNA biosensors with spatiotemporal precision. We also provide recent examples for improvements made to the caging groups and demonstrate their impact in practical applications such as imaging of metal ions. Finally, current limitations and potential opportunities for future photocontrollable FNA sensors are also discussed.
Section snippets
Photocontrol of catalytic nucleic acids based on photolabile modifications
Photolabile modification is a frequently used caging method for controlling the configuration and activity of nucleic acids. These photolabile modifications include photoresponsive small molecule caging of a single nucleotide and photocleavable (PC) linkers between two nucleotides. Detailed properties and development of these photolabile groups in controlling DNA nanomaterials have been reviewed elsewhere [∗∗22], and we will focus on the application of these photolabile groups in functional
Photocontrol of aptamers based on photolabile modifications
Photolabile groups have also been used in regulating the functions of aptamers by blocking the binding interface for target recognition (Figure 5a). For example, incorporating photocaged thymidine phosphoramidites into the target-binding site of the thrombin binding aptamer (TBA) prevented thrombin from binding, and the binding can be restored by using 366 nm light to remove the photocage group [45]. Instead of blocking the binding domain of the aptamers using a photocage group, other studies
Summary and future directions
In this review, we have summarized recent progress in spatiotemporal control of photocaged functional nucleic acids for sensing and imaging in biology by discussing methods to control the functions of catalytic nucleic acids and aptamers based on photolabile modifications, photoisomerization, and, in the case of DNAzymes, photothermal activation. While many uncaging methods have been reported, the efficiency is still relatively low. Developing novel photolabile and photoswitching groups that
Declaration of Competing Interest
Nothing declared.
Acknowledgements
The authors thank the Lu group members and collaborators who have contributed over the years to the many studies described here, Gregory Pawel and Ryan Lake for proofreading, and the U.S. National Institutes of Health (GM124316 and MH110975) and DOE Center for Advanced Bioenergy and Bioproducts Innovation (U.S. Department of Energy, Office of Science, Office of Biological and Environmental Research under Award Number DE-SC0018420) for financial support. Zhenglin Yang acknowledges support from
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