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Stable tri-snRNP integration is accompanied by a major structural rearrangement of the spliceosome that is dependent on Prp8 interaction with the 5‘ splice site.

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

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Rigo,  N.
Department of Cellular Biochemistry, MPI for biophysical chemistry, Max Planck Society;

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Agafonov,  D. E.
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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Urlaub,  H.
Research Group of Bioanalytical Mass Spectrometry, MPI for biophysical chemistry, Max Planck Society;

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Will,  C. L.
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;

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Citation

Boesler, C., Rigo, N., Agafonov, D. E., Kastner, B., Urlaub, H., Will, C. L., et al. (2015). Stable tri-snRNP integration is accompanied by a major structural rearrangement of the spliceosome that is dependent on Prp8 interaction with the 5‘ splice site. RNA, 21(11), 1993-2005. doi:10.1261/rna.053991.115.


Cite as: https://hdl.handle.net/11858/00-001M-0000-0028-4609-E
Abstract
Exon definition is the predominant initial spliceosome assembly pathway in higher eukaryotes, but it remains much less well-characterized compared to the intron-defined assembly pathway. Addition in trans of an excess of 5′ss containing RNA to a splicing reaction converts a 37S exon-defined complex, formed on a single exon RNA substrate, into a 45S B-like spliceosomal complex with stably integrated U4/U6.U5 tri-snRNP. This 45S complex is compositonally and structurally highly similar to an intron-defined spliceosomal B complex. Stable tri-snRNP integration during B-like complex formation is accompanied by a major structural change as visualized by electron microscopy. The changes in structure and stability during transition from a 37S to 45S complex can be induced in affinity-purified cross-exon complexes by adding solely the 5′ss RNA oligonucleotide. This conformational change does not require the B-specific proteins, which are recruited during this stabilization process, or site-specific phosphorylation of hPrp31. Instead it is triggered by the interaction of U4/U6.U5 tri-snRNP components with the 5′ss sequence, most importantly between Prp8 and nucleotides at the exon–intron junction. These studies provide novel insights into the conversion of a cross-exon to cross-intron organized spliceosome and also shed light on the requirements for stable tri-snRNP integration during B complex formation.