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Hierarchical, clustered protein interact ions with U4/U6 snRNA: a biochemical role for U4/U6 proteins

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Nottrott,  S.
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;

Luehrmann,  R.
Max Planck Society;

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Citation

Nottrott, S., Urlaub, H., & Luehrmann, R. (2002). Hierarchical, clustered protein interact ions with U4/U6 snRNA: a biochemical role for U4/U6 proteins. EMBO Journal, 21(20), 5527-5538. Retrieved from http://www.nature.com/emboj/journal/v21/n20/pdf/7594761a.pdf.


Cite as: http://hdl.handle.net/11858/00-001M-0000-0012-F2C2-2
Abstract
During activation of the spliceosome,, the U4/U6 snRNA duplex is dissociated, releasing U6 for subsequent base pairing with U2 snRNA. Proteins that directly bind the U4/U6 interaction domain potentially could mediate these structural changes. We thus investigated binding of the human U4/U6-specific proteins, 15.5K, 61K and the 20/60/90K protein complex, to U4/U6 snRNA in vitro. We demonstrate that protein 15.5K is a nucleation factor for U4/U6 snRNP assembly, mediating the interaction of 61K and 20/60/90K with U4/U6 snRNA. A similar hierarchical assembly pathway is observed for the U4atac/U6atac snRNP. In addition, we show that protein 61K directly contacts the 5' portion of U4 snRNA via a novel RNA-binding domain. Furthermore, the 20/60/90K heteromer requires stem II but not stem I of the U4/U6 duplex for binding, and this interaction involves a direct contact between protein 90K and U6. This uneven clustering of the U4/U6 snRNP-specific proteins on U4/U6 snRNA is consistent with a sequential dissociation of the U4/U6 duplex prior to spliceosome catalysis.