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Journal Article

Retention of posttranscriptional spliceosomes in nuclear speckles until splicing completion.

MPS-Authors
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Will,  C. L.
Department of Cellular Biochemistry, MPI for biophysical chemistry, Max Planck Society;

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Peng,  J.
Department of Cellular Biochemistry, MPI for biophysical chemistry, Max Planck Society;

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Makarov,  E. M.
Department of Cellular Biochemistry, MPI for biophysical chemistry, Max Planck Society;

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Kastner,  B.
Department of Cellular Biochemistry, MPI for biophysical chemistry, Max Planck Society;

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Lemm,  I.
Department of Cellular Biochemistry, MPI for biophysical chemistry, Max Planck Society;

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Urlaub,  Henning
Research Group of Bioanalytical Mass Spectrometry, MPI for biophysical chemistry, Max Planck Society;

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Hartmuth,  K.
Department of Cellular Biochemistry, MPI for biophysical chemistry, Max Planck Society;

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Lührmann,  R.
Department of Cellular Biochemistry, MPI for biophysical chemistry, Max Planck Society;

Fulltext (public)

1481223.pdf
(Publisher version), 2MB

Supplementary Material (public)

1481223_Supplement_1.pdf
(Supplementary material), 9MB

Citation

Girard, C., Will, C. L., Peng, J., Makarov, E. M., Kastner, B., Lemm, I., et al. (2012). Retention of posttranscriptional spliceosomes in nuclear speckles until splicing completion. Nature communications, 3: 994. doi:10.1038/ncomms1998.


Cite as: http://hdl.handle.net/11858/00-001M-0000-000F-A53B-9
Abstract
There is little quantitative information regarding how much splicing occurs co-transcriptionally in higher eukaryotes, and it remains unclear where precisely splicing occurs in the nucleus. Here we determine the global extent of co- and post-transcriptional splicing in mammalian cells, and their respective subnuclear locations, using antibodies that specifically recognize phosphorylated SF3b155 (P-SF3b155) found only in catalytically activated/active spliceosomes. Quantification of chromatin- and nucleoplasm-associated P-SF3b155 after fractionation of HeLa cell nuclei, reveals that ~80% of pre-mRNA splicing occurs co-transcriptionally. Active spliceosomes localize in situ to regions of decompacted chromatin, at the periphery of or within nuclear speckles. Immunofluorescence microscopy with anti-P-SF3b155 antibodies, coupled with transcription inhibition and a block in splicing after SF3b155 phosphorylation, indicates that post-transcriptional splicing occurs in nuclear speckles and that release of post-transcriptionally spliced mRNA from speckles is coupled to the nuclear mRNA export pathway. Our data provide new insights into when and where splicing occurs in cells.