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The differential interaction of snRNPs with pre-mRNA reveals splicing kinetics in living cells.

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Poser,  Ina
Max Planck Institute of Molecular Cell Biology and Genetics, Max Planck Society;

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Neugebauer,  Karla M.
Max Planck Institute of Molecular Cell Biology and Genetics, Max Planck Society;

Stanĕk,  David
Max Planck Society;

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Citation

Huranová, M., Ivani, I., Benda, A., Poser, I., Brody, Y., Hof, M., et al. (2010). The differential interaction of snRNPs with pre-mRNA reveals splicing kinetics in living cells. The Journal of Cell Biology, 191(1), 75-86.


Cite as: http://hdl.handle.net/21.11116/0000-0001-0C45-5
Abstract
Precursor messenger RNA (pre-mRNA) splicing is catalyzed by the spliceosome, a large ribonucleoprotein (RNP) complex composed of five small nuclear RNP particles (snRNPs) and additional proteins. Using live cell imaging of GFP-tagged snRNP components expressed at endogenous levels, we examined how the spliceosome assembles in vivo. A comprehensive analysis of snRNP dynamics in the cell nucleus enabled us to determine snRNP diffusion throughout the nucleoplasm as well as the interaction rates of individual snRNPs with pre-mRNA. Core components of the spliceosome, U2 and U5 snRNPs, associated with pre-mRNA for 15-30 s, indicating that splicing is accomplished within this time period. Additionally, binding of U1 and U4/U6 snRNPs with pre-mRNA occurred within seconds, indicating that the interaction of individual snRNPs with pre-mRNA is distinct. These results are consistent with the predictions of the step-wise model of spliceosome assembly and provide an estimate on the rate of splicing in human cells.