Abstract
The replication of a virus within its host cell involves numerous interactions between viral and cellular factors, which have to be tightly controlled in space and time. The intricate interplay between viral exploitation of cellular pathways and the intrinsic host defense mechanisms is difficult to unravel by traditional bulk approaches. In recent years, novel fluorescence microscopy techniques and single virus tracking have transformed the investigation of dynamic virus-host interactions. A prerequisite for the application of these imaging-based methods is the attachment of a fluorescent label to the structure of interest. However, their small size, limited coding capacity and multifunctional proteins render viruses particularly challenging targets for fluorescent labeling approaches. Click chemistry in conjunction with genetic code expansion provides virologists with a novel toolbox for site-specific, minimally invasive labeling of virion components, whose potential has just recently begun to be exploited. Here, we summarize recent achievements, current developments and future challenges for the labeling of viral nucleic acids, proteins, glycoproteins or lipids using click chemistry in order to study dynamic processes in virus-cell interactions.
Highlights
Viruses occupy a unique position at the boundary of life
Advanced fluorescence microscopy methods have specific requirements regarding the photophysical properties of the fluorophore, which are often not met by fluorescent proteins (FPs)
We summarize concepts, current achievements and future challenges in the use of click chemistry for investigating virus-host cell interactions by advanced imaging approaches
Summary
Viruses occupy a unique position at the boundary of life. Outside of their host, they exist as (largely) inert macromolecular assemblies, which awaken to a “borrowed life” upon entry into a suitable host cell. For the labeling of proteins, the genetically encoded marker green fluorescent protein (GFP) and the constantly expanding panel of engineered fluorescent proteins (FPs) [7] have been, and still are, invaluable tools for the investigation of dynamic events in cell biology Besides their relatively high molecular mass of ~ 27 kDa, FPs have clear limitations regarding the use of advanced light microscopic methods and the study of viruses in general [8]. Advanced fluorescence microscopy methods have specific requirements regarding the photophysical properties of the fluorophore, which are often not met by FPs. Single virus tracking within an infected cell requires high fluorescence intensity of the label for sensitive and quantitative detection of individual particles, and high photostability of the dye to allow for the acquisition of image sequences with high temporal resolution. We summarize concepts, current achievements and future challenges in the use of click chemistry for investigating virus-host cell interactions by advanced imaging approaches
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