This study reports an experimental system that allows the visualization of transcription and mRNA splicing in living human cells, with single-molecule resolution. Using this approach, the authors obtained key insights into splicing dynamics. First, they inserted GFP-tagged versions of the coat protein of bacteriophage MS2 or the antiterminator protein N of bacteriophage λ into each of the two introns of the β-globin gene and followed splicing dynamics using microscopy. They observed that β-globin intron excision occurs in 20–30 seconds, with the first intron having a shorter lifetime than the second (terminal) intron, which suggests that it is excised while the terminal intron is still present. Moreover, using a mouse immunoglobulin M (IgM) gene engineered to contain one intron between exons M1 and M2, the authors showed that splice site strength also influences splicing kinetics, as introns with a strong polypyrimidine tract (Py tract; an essential splicing signal) had a shorter lifetime than those with a weak Py tract.
References
Martin, R. M. et al. Live-cell visualization of pre-mRNA splicing with single-molecule sensitivity. Cell Rep. http://dx.doi.org/10.1016/j.celrep.2013.08.013 (2013)
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David, R. Visualizing RNA splicing. Nat Rev Mol Cell Biol 14, 688 (2013). https://doi.org/10.1038/nrm3689
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DOI: https://doi.org/10.1038/nrm3689