TY - JOUR
T1 - The differential interaction of snRNPs with pre-mRNA reveals splicing kinetics in living cells
AU - Huranová, Martina
AU - Ivani, Ivan
AU - Benda, Aleš
AU - Poser, Ina
AU - Brody, Yehuda
AU - Hof, Martin
AU - Shav-Tal, Yaron
AU - Neugebauer, Karla M.
AU - Staněk, David
PY - 2010/10/4
Y1 - 2010/10/4
N2 - Precursor messenger RNA (pre-mRNA) splicing is catalyzed by the spliceosome, a large ribonucleo-protein (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.
AB - Precursor messenger RNA (pre-mRNA) splicing is catalyzed by the spliceosome, a large ribonucleo-protein (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.
UR - http://www.scopus.com/inward/record.url?scp=77957747157&partnerID=8YFLogxK
U2 - 10.1083/jcb.201004030
DO - 10.1083/jcb.201004030
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C2 - 20921136
AN - SCOPUS:77957747157
SN - 0021-9525
VL - 191
SP - 75
EP - 86
JO - Journal of Cell Biology
JF - Journal of Cell Biology
IS - 1
ER -