Structural basis for dual roles of Aar2p in U5 snRNP assembly
- Gert Weber1,4,
- Vanessa F. Cristão2,
- Karine F. Santos1,
- Sina Mozaffari Jovin3,
- Anna C. Heroven1,
- Nicole Holton1,
- Reinhard Lührmann3,
- Jean D. Beggs2 and
- Markus C. Wahl1,4
- 1Fachbereich Biologie/Chemie/Pharmazie, Abteilung Strukturbiochemie, Freie Universität Berlin, D-14195 Berlin, Germany;
- 2Wellcome Trust Centre for Cell Biology, University of Edinburgh, Edinburgh EH9 3JR, United Kingdom;
- 3Zelluläre Biochemie, Max-Planck-Institut für Biophysikalische Chemie, D-37077 Göttingen, Germany
Abstract
Yeast U5 small nuclear ribonucleoprotein particle (snRNP) is assembled via a cytoplasmic precursor that contains the U5-specific Prp8 protein but lacks the U5-specific Brr2 helicase. Instead, pre-U5 snRNP includes the Aar2 protein not found in mature U5 snRNP or spliceosomes. Aar2p and Brr2p bind competitively to a C-terminal region of Prp8p that comprises consecutive RNase H-like and Jab1/MPN-like domains. To elucidate the molecular basis for this competition, we determined the crystal structure of Aar2p in complex with the Prp8p RNase H and Jab1/MPN domains. Aar2p binds on one side of the RNase H domain and extends its C terminus to the other side, where the Jab1/MPN domain is docked onto a composite Aar2p–RNase H platform. Known Brr2p interaction sites of the Jab1/MPN domain remain available, suggesting that Aar2p-mediated compaction of the Prp8p domains sterically interferes with Brr2p binding. Moreover, Aar2p occupies known RNA-binding sites of the RNase H domain, and Aar2p interferes with binding of U4/U6 di-snRNA to the Prp8p C-terminal region. Structural and functional analyses of phospho-mimetic mutations reveal how phosphorylation reduces affinity of Aar2p for Prp8p and allows Brr2p and U4/U6 binding. Our results show how Aar2p regulates both protein and RNA binding to Prp8p during U5 snRNP assembly.
Keywords
- assembly chaperone
- pre-mRNA splicing
- regulation by phosphorylation
- snRNP biogenesis and recycling
- X-ray crystallography
Footnotes
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↵4 Corresponding authors
E-mail mwahl{at}zedat.fu-berlin.de
E-mail gweber{at}chemie.fu-berlin.de
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Supplemental material is available for this article.
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Article published online ahead of print. Article and publication date are online at http://www.genesdev.org/cgi/doi/10.1101/gad.213207.113.
- Received January 3, 2013.
- Accepted February 1, 2013.
- Copyright © 2013 by Cold Spring Harbor Laboratory Press