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Post-transcriptional spliceosomes are retained in nuclear speckles until splicing completion
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Post-transcriptional spliceosomes are retained in nuclear speckles until splicing completion
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Journal Article
Post-transcriptional spliceosomes are retained in nuclear speckles until splicing completion
2012
Overview
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.
It is unclear where in the nucleus splicing takes place and how much occurs post-transcriptionally. Using antibodies raised against a phosphorylated splicing factor, Girard
et al
. show that the majority of splicing occurs co-transcriptionally and that post-transcriptional splicing occurs in nuclear speckles.
Publisher
Nature Publishing Group UK,Nature Publishing Group
