The Dynamics of mRNA Turnover Revealed by Single-Molecule Imaging in Single Cells

摘要

Summary RNA degradation plays a fundamental role in regulating gene expression. In order to characterize the spatiotemporal dynamics of RNA turnover in single cells, we developed a fluorescent biosensor based on dual-color, single-molecule RNA imaging that allows intact transcripts to be distinguished from stabilized degradation intermediates. Using this method, we measured mRNA decay in single cells and found that individual degradation events occur independently within the cytosol and are not enriched within processing bodies. We show that slicing of an mRNA targeted for endonucleolytic cleavage by the RNA-induced silencing complex can be observed in real time in living cells. This methodology provides a framework for investigating the entire life history of individual mRNAs from birth to death in single cells. Graphical Abstract Display Omitted Highlights ? TREAT enables single-mRNA imaging of RNA degradation in single cells ? TREAT mRNA degradation does not burst ? Real-time observation of Ago2 slicing of TREAT mRNAs ? TREAT mRNAs are not degraded in P-bodies RNA degradation plays a fundamental role in regulating gene expression. Horvathova et?al. developed a fluorescence microscopy methodology that enables the dynamics of RNA degradation to be quantified with single-molecule resolution in fixed and living cells. This allows the entire life of mRNAs from birth to death to be visualized.