Abstract
The ASAP complex interacts with the exon-junction complex (EJC), a messenger ribonucleoprotein complex involved in post-transcriptional regulation. The three ASAP subunits (Acinus, RNPS1 and SAP18) have been individually implicated in transcriptional regulation, pre-mRNA splicing and mRNA quality control. To shed light on the basis for and consequences of ASAP's interaction with the EJC, we have determined the 1.9-Å resolution structure of a eukaryotic ASAP core complex. The RNA-recognition motif of RNPS1 binds to a conserved motif of Acinus with a recognition mode similar to that observed in splicing U2AF proteins. The Acinus–RNPS1 platform recruits the ubiquitin-like domain of SAP18, forming a ternary complex that has both RNA- and protein-binding properties. Unexpectedly, our structural analysis identified an Acinus-like motif in Pinin, another EJC-associated splicing factor. We show that Pinin physically interacts with RNPS1 and SAP18, forming an alternative ternary complex, PSAP.
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Acknowledgements
We would like to thank J. Basquin, K. Valer-Saldaña and S. Pleyer at the Max Planck Institute Biochemistry Crystallization Facility; the Max Planck Institute Biochemistry Core Facility for mass spectrometry; F. Bonneau for help with immunoprecipitation and RNA-binding assays; M. Jinek (University of California, Berkeley) for the HA and HA-Flag vectors ; M. Rode and C. Tommasetto for baculovirus expression; J. Söding for help with the bioinformatic search of RSB-containing proteins; D.L. Makino with help on data collection; and A. Pauluhn and the staff at the Swiss Light Source synchrotron. We also would like to thank the members of our labs for discussions and for critical reading of the manuscript.
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Author contributions to this work were as follows: purification, structure determination, biochemical assays and computational analysis were done by A.G.M.; ASAP limited proteolysis, crystallization and data collection by J.E.; fluorescence anisotropy shown in Figures 3 and 4 by C.B.; and the pull-down assay shown in Supplementary Figure 1 was done by H.L.H. E.C. supervised the project. E.C. and A.G.M. wrote the paper.
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Murachelli, A., Ebert, J., Basquin, C. et al. The structure of the ASAP core complex reveals the existence of a Pinin-containing PSAP complex. Nat Struct Mol Biol 19, 378–386 (2012). https://doi.org/10.1038/nsmb.2242
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DOI: https://doi.org/10.1038/nsmb.2242
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