Cryo-EM structure of the spliceosome immediately after branching (original) (raw)

• Will, C. L. & Lührmann, R. Spliceosome structure and function. Cold Spring Harb. Perspect. Biol. 3, a003707 (2011)
  CAS PubMed PubMed Central Google Scholar
• Burge, C. B., Tuschl, T. & Sharp, P. A. in The RNA World II (eds Gesteland, R. F., Cech, T. R. & Atkins, J. F. ) 525–560 (Cold Spring Harbor Laboratory Press, 1999)
• Lambowitz, A. M. & Zimmerly, S. Group II introns: mobile ribozymes that invade DNA. Cold Spring Harb. Perspect. Biol. 3, a003616 (2011)
  PubMed PubMed Central Google Scholar
• Chan, S.-P., Kao, D.-I., Tsai, W.-Y. & Cheng, S.-C. The Prp19p-associated complex in spliceosome activation. Science 302, 279–282 (2003)
  ADS CAS PubMed Google Scholar
• Ohi, M. D. & Gould, K. L. Characterization of interactions among the Cef1p-Prp19p-associated splicing complex. RNA 8, 798–815 (2002)
  CAS PubMed PubMed Central Google Scholar
• Fabrizio, P. et al. The evolutionarily conserved core design of the catalytic activation step of the yeast spliceosome. Mol. Cell 36, 593–608 (2009)
  CAS PubMed Google Scholar
• Lesser, C. F. & Guthrie, C. Mutations in U6 snRNA that alter splice site specificity: implications for the active site. Science 262, 1982–1988 (1993)
  ADS CAS PubMed Google Scholar
• Kandels-Lewis, S. & Séraphin, B. Involvement of U6 snRNA in 5′ splice site selection. Science 262, 2035–2039 (1993)
  ADS CAS PubMed Google Scholar
• Raghunathan, P. L. & Guthrie, C. RNA unwinding in U4/U6 snRNPs requires ATP hydrolysis and the DEIH-box splicing factor Brr2. Curr. Biol. 8, 847–855 (1998)
  CAS PubMed Google Scholar
• Laggerbauer, B., Achsel, T. & Lührmann, R. The human U5-200kD DEXH-box protein unwinds U4/U6 RNA duplices in vitro. Proc. Natl Acad. Sci. USA 95, 4188–4192 (1998)
  ADS CAS PubMed PubMed Central Google Scholar
• Newman, A. J. & Norman, C. U5 snRNA interacts with exon sequences at 5′ and 3′ splice sites. Cell 68, 743–754 (1992)
  CAS PubMed Google Scholar
• Sontheimer, E. J. & Steitz, J. A. The U5 and U6 small nuclear RNAs as active site components of the spliceosome. Science 262, 1989–1996 (1993)
  ADS CAS PubMed Google Scholar
• Ohrt, T. et al. Molecular dissection of step 2 catalysis of yeast pre-mRNA splicing investigated in a purified system. RNA 19, 902–915 (2013)
  CAS PubMed PubMed Central Google Scholar
• Cordin, O., Hahn, D. & Beggs, J. D. Structure, function and regulation of spliceosomal RNA helicases. Curr. Opin. Cell Biol. 24, 431–438 (2012)
  CAS PubMed Google Scholar
• Schwer, B. A conformational rearrangement in the spliceosome sets the stage for Prp22-dependent mRNA release. Mol. Cell 30, 743–754 (2008)
  CAS PubMed PubMed Central Google Scholar
• Company, M., Arenas, J. & Abelson, J. Requirement of the RNA helicase-like protein PRP22 for release of messenger RNA from spliceosomes. Nature 349, 487–493 (1991)
  ADS CAS PubMed Google Scholar
• Tsai, R.-T. et al. Spliceosome disassembly catalyzed by Prp43 and its associated components Ntr1 and Ntr2. Genes Dev. 19, 2991–3003 (2005)
  CAS PubMed PubMed Central Google Scholar
• Abelson, J. et al. Conformational dynamics of single pre-mRNA molecules during in vitro splicing. Nat. Struct. Mol. Biol. 17, 504–512 (2010)
  CAS PubMed PubMed Central Google Scholar
• Nguyen, T. H. D. et al. Cryo-EM structure of the yeast U4/U6.U5 tri-snRNP at 3.7 Å resolution. Nature 530, 298–302 (2016)
  ADS CAS PubMed PubMed Central Google Scholar
• Wan, R. et al. The 3.8 Å structure of the U4/U6.U5 tri-snRNP: Insights into spliceosome assembly and catalysis. Science 351, 466–475 (2016)
  ADS CAS PubMed Google Scholar
• Galej, W. P., Oubridge, C., Newman, A. J. & Nagai, K. Crystal structure of Prp8 reveals active site cavity of the spliceosome. Nature 493, 638–643 (2013)
  ADS CAS PubMed PubMed Central Google Scholar
• Rasche, N. et al. Cwc2 and its human homologue RBM22 promote an active conformation of the spliceosome catalytic centre. EMBO J. 31, 1591–1604 (2012)
  CAS PubMed PubMed Central Google Scholar
• Hilliker, A. K., Mefford, M. A. & Staley, J. P. U2 toggles iteratively between the stem IIa and stem IIc conformations to promote pre-mRNA splicing. Genes Dev. 21, 821–834 (2007)
  CAS PubMed PubMed Central Google Scholar
• Perriman, R. J. & Ares, M., Jr. Rearrangement of competing U2 RNA helices within the spliceosome promotes multiple steps in splicing. Genes Dev. 21, 811–820 (2007)
  CAS PubMed PubMed Central Google Scholar
• Grainger, R. J., Barrass, J. D., Jacquier, A., Rain, J.-C. & Beggs, J. D. Physical and genetic interactions of yeast Cwc21p, an ortholog of human SRm300/SRRM2, suggest a role at the catalytic center of the spliceosome. RNA 15, 2161–2173 (2009)
  CAS PubMed PubMed Central Google Scholar
• Yan, C. et al. Structure of a yeast spliceosome at 3.6-angstrom resolution. Science 349, 1182–1191 (2015)
  ADS CAS PubMed Google Scholar
• Nguyen, T. H. D. et al. Structural basis of Brr2-Prp8 interactions and implications for U5 snRNP biogenesis and the spliceosome active site. Structure 21, 910–919 (2013)
  CAS PubMed PubMed Central Google Scholar
• van Nues, R. W. & Beggs, J. D. Functional contacts with a range of splicing proteins suggest a central role for Brr2p in the dynamic control of the order of events in spliceosomes of Saccharomyces cerevisiae. Genetics 157, 1451–1467 (2001)
  CAS PubMed PubMed Central Google Scholar
• Smith, D. J., Query, C. C. & Konarska, M. M. “Nought may endure but mutability”: spliceosome dynamics and the regulation of splicing. Mol. Cell 30, 657–666 (2008)
  CAS PubMed PubMed Central Google Scholar
• Konarska, M. M., Vilardell, J. & Query, C. C. Repositioning of the reaction intermediate within the catalytic center of the spliceosome. Mol. Cell 21, 543–553 (2006)
  CAS PubMed Google Scholar
• Kim, C. H. & Abelson, J. Site-specific crosslinks of yeast U6 snRNA to the pre-mRNA near the 5′ splice site. RNA 2, 995–1010 (1996)
  CAS PubMed PubMed Central Google Scholar
• Buchwald, G., Schüssler, S., Basquin, C., Le Hir, H. & Conti, E. Crystal structure of the human eIF4AIII-CWC22 complex shows how a DEAD-box protein is inhibited by a MIF4G domain. Proc. Natl Acad. Sci. USA 110, E4611–E4618 (2013)
  ADS CAS PubMed PubMed Central Google Scholar
• Madhani, H. D. & Guthrie, C. A novel base-pairing interaction between U2 and U6 snRNAs suggests a mechanism for the catalytic activation of the spliceosome. Cell 71, 803–817 (1992)
  CAS PubMed Google Scholar
• Fica, S. M., Mefford, M. A., Piccirilli, J. A. & Staley, J. P. Evidence for a group II intron-like catalytic triplex in the spliceosome. Nat. Struct. Mol. Biol. 21, 464–471 (2014)
  CAS PubMed PubMed Central Google Scholar
• Steitz, T. A. & Steitz, J. A. A general two-metal-ion mechanism for catalytic RNA. Proc. Natl Acad. Sci. USA 90, 6498–6502 (1993)
  ADS CAS PubMed MATH PubMed Central Google Scholar
• Fica, S. M. et al. RNA catalyses nuclear pre-mRNA splicing. Nature 503, 229–234 (2013)
  ADS CAS PubMed PubMed Central Google Scholar
• Robart, A. R., Chan, R. T., Peters, J. K., Rajashankar, K. R. & Toor, N. Crystal structure of a eukaryotic group II intron lariat. Nature 514, 193–197 (2014)
  ADS CAS PubMed PubMed Central Google Scholar
• Chen, H.-C., Tseng, C.-K., Tsai, R.-T., Chung, C.-S. & Cheng, S.-C. Link of NTR-mediated spliceosome disassembly with DEAH-box ATPases Prp2, Prp16, and Prp22. Mol. Cell. Biol. 33, 514–525 (2013)
  CAS PubMed PubMed Central Google Scholar
• Warkocki, Z. et al. Reconstitution of both steps of Saccharomyces cerevisiae splicing with purified spliceosomal components. Nat. Struct. Mol. Biol. 16, 1237–1243 (2009)
  CAS PubMed Google Scholar
• Chiu, Y.-F. et al. Cwc25 is a novel splicing factor required after Prp2 and Yju2 to facilitate the first catalytic reaction. Mol. Cell. Biol. 29, 5671–5678 (2009)
  CAS PubMed PubMed Central Google Scholar
• Krishnan, R. et al. Biased Brownian ratcheting leads to pre-mRNA remodeling and capture prior to first-step splicing. Nat. Struct. Mol. Biol. 20, 1450–1457 (2013)
  CAS PubMed PubMed Central Google Scholar
• Voorhees, R. M., Weixlbaumer, A., Loakes, D., Kelley, A. C. & Ramakrishnan, V. Insights into substrate stabilization from snapshots of the peptidyl transferase center of the intact 70S ribosome. Nat. Struct. Mol. Biol. 16, 528–533 (2009)
  CAS PubMed PubMed Central Google Scholar
• Semlow, D. R., Blanco, M. R., Walter, N. G. & Staley, J. P. Spliceosomal DEAH-Box ATPases remodel pre-mRNA to activate alternative splice sites. Cell 164, 985–998 (2016)
  CAS PubMed PubMed Central Google Scholar
• Tseng, C.-K., Liu, H.-L. & Cheng, S.-C. DEAH-box ATPase Prp16 has dual roles in remodeling of the spliceosome in catalytic steps. RNA 17, 145–154 (2011)
  CAS PubMed PubMed Central Google Scholar
• Schwer, B. & Guthrie, C. A conformational rearrangement in the spliceosome is dependent on PRP16 and ATP hydrolysis. EMBO J. 11, 5033–5039 (1992)
  CAS PubMed PubMed Central Google Scholar
• Villa, T. & Guthrie, C. The Isy1p component of the NineTeen complex interacts with the ATPase Prp16p to regulate the fidelity of pre-mRNA splicing. Genes Dev. 19, 1894–1904 (2005)
  CAS PubMed PubMed Central Google Scholar
• Umen, J. G. & Guthrie, C. Prp16p, Slu7p, and Prp8p interact with the 3′ splice site in two distinct stages during the second catalytic step of pre-mRNA splicing. RNA 1, 584–597 (1995)
  CAS PubMed PubMed Central Google Scholar
• Chen, W. et al. Endogenous U2·U5·U6 snRNA complexes in S. pombe are intron lariat spliceosomes. RNA 20, 308–320 (2014)
  CAS PubMed PubMed Central Google Scholar
• Nguyen, T. H. D. et al. CryoEM structures of two spliceosomal complexes: starter and dessert at the spliceosome feast. Curr. Opin. Struct. Biol. 36, 48–57 (2016)
  CAS PubMed PubMed Central Google Scholar
• Garrey, S. M. et al. A homolog of lariat-debranching enzyme modulates turnover of branched RNA. RNA 20, 1337–1348 (2014)
  CAS PubMed PubMed Central Google Scholar
• Schreieck, A. et al. RNA polymerase II termination involves C-terminal-domain tyrosine dephosphorylation by CPF subunit Glc7. Nat. Struct. Mol. Biol. 21, 175–179 (2014)
  CAS PubMed PubMed Central Google Scholar
• Lin, R. J., Newman, A. J., Cheng, S. C. & Abelson, J. Yeast mRNA splicing in vitro. J. Biol. Chem. 260, 14780–14792 (1985)
  CAS PubMed Google Scholar
• Zhou, Z., Licklider, L. J., Gygi, S. P. & Reed, R. Comprehensive proteomic analysis of the human spliceosome. Nature 419, 182–185 (2002)
  ADS CAS PubMed Google Scholar
• Hardin, J. W., Warnasooriya, C., Kondo, Y., Nagai, K. & Rueda, D. Assembly and dynamics of the U4/U6 di-snRNP by single-molecule FRET. Nucleic Acids Res. 43, 10963–10974 (2015)
  CAS PubMed PubMed Central Google Scholar
• Nguyen, T. H. D. et al. The architecture of the spliceosomal U4/U6.U5 tri-snRNP. Nature 523, 47–52 (2015)
  ADS CAS PubMed PubMed Central Google Scholar
• Li, X. et al. Electron counting and beam-induced motion correction enable near-atomic-resolution single-particle cryo-EM. Nat. Methods 10, 584–590 (2013)
  CAS PubMed PubMed Central Google Scholar
• Rohou, A. & Grigorieff, N. CTFFIND4: Fast and accurate defocus estimation from electron micrographs. J. Struct. Biol. 192, 216–221 (2015)
  PubMed PubMed Central Google Scholar
• Scheres, S. H. W. RELION: implementation of a Bayesian approach to cryo-EM structure determination. J. Struct. Biol. 180, 519–530 (2012)
  CAS PubMed PubMed Central Google Scholar
• Scheres, S. H. W. Semi-automated selection of cryo-EM particles in RELION-1.3. J. Struct. Biol. 189, 114–122 (2015)
  ADS CAS PubMed PubMed Central Google Scholar
• Bai, X.-C., Rajendra, E., Yang, G., Shi, Y. & Scheres, S. H. W. Sampling the conformational space of the catalytic subunit of human γ-secretase . eLife 4, 1485 (2015)
  Google Scholar
• Scheres, S. H. W. & Chen, S. Prevention of overfitting in cryo-EM structure determination. Nat. Methods 9, 853–854 (2012)
  CAS PubMed PubMed Central Google Scholar
• Chen, S. et al. High-resolution noise substitution to measure overfitting and validate resolution in 3D structure determination by single particle electron cryomicroscopy. Ultramicroscopy 135, 24–35 (2013)
  CAS PubMed PubMed Central Google Scholar
• Kucukelbir, A., Sigworth, F. J. & Tagare, H. D. Quantifying the local resolution of cryo-EM density maps. Nat. Methods 11, 63–65 (2014)
  CAS PubMed Google Scholar
• Lu, P. et al. Structure of the mRNA splicing complex component Cwc2: insights into RNA recognition. Biochem. J. 441, 591–597 (2012)
  CAS PubMed Google Scholar
• van Roon, A.-M. M. et al. 113Cd NMR experiments reveal an unusual metal cluster in the solution structure of the yeast splicing protein Bud31p. Angew. Chem. Int. Edn Engl. 54, 4861–4864 (2015)
  CAS Google Scholar
• Biasini, M. et al. SWISS-MODEL: modelling protein tertiary and quaternary structure using evolutionary information. Nucleic Acids Res. 42, W252–W258 (2014)
  CAS PubMed PubMed Central Google Scholar
• Kurowski, M. A. & Bujnicki, J. M. GeneSilico protein structure prediction meta-server. Nucleic Acids Res. 31, 3305–3307 (2003)
  CAS PubMed PubMed Central Google Scholar
• Price, S. R., Evans, P. R. & Nagai, K. Crystal structure of the spliceosomal U2B"-U2A′ protein complex bound to a fragment of U2 small nuclear RNA. Nature 394, 645–650 (1998)
  ADS CAS PubMed Google Scholar
• Kudla, G., Granneman, S., Hahn, D., Beggs, J. D. & Tollervey, D. Cross-linking, ligation, and sequencing of hybrids reveals RNA-RNA interactions in yeast. Proc. Natl Acad. Sci. USA 108, 10010–10015 (2011)
  ADS CAS PubMed PubMed Central Google Scholar
• Yang, J. et al. The I-TASSER Suite: protein structure and function prediction. Nat. Methods 12, 7–8 (2015)
  CAS PubMed PubMed Central Google Scholar
• Walbott, H. et al. Prp43p contains a processive helicase structural architecture with a specific regulatory domain. EMBO J. 29, 2194–2204 (2010)
  CAS PubMed PubMed Central Google Scholar
• Murshudov, G. N., Vagin, A. A. & Dodson, E. J. Refinement of macromolecular structures by the maximum-likelihood method. Acta Crystallogr. D Biol. Crystallogr. 53, 240–255 (1997)
  CAS PubMed Google Scholar
• Nicholls, R. A., Fischer, M., McNicholas, S. & Murshudov, G. N. Conformation-independent structural comparison of macromolecules with ProSMART. Acta Crystallogr. D Biol. Crystallogr. 70, 2487–2499 (2014)
  CAS PubMed PubMed Central Google Scholar
• Brown, A. et al. Tools for macromolecular model building and refinement into electron cryo-microscopy reconstructions. Acta Crystallogr. D Biol. Crystallogr. 71, 136–153 (2015)
  CAS PubMed PubMed Central Google Scholar
• Goddard, T. D., Huang, C. C. & Ferrin, T. E. Visualizing density maps with UCSF Chimera. J. Struct. Biol. 157, 281–287 (2007)
  CAS PubMed Google Scholar
• Marcia, M. & Pyle, A. M. Visualizing group II intron catalysis through the stages of splicing. Cell 151, 497–507 (2012)
  CAS PubMed PubMed Central Google Scholar
• Le Hir, H., Saulière, J. & Wang, Z. The exon junction complex as a node of post-transcriptional networks. Nat. Rev. Mol. Cell Biol. 17, 41–54 (2016)
  PubMed Google Scholar
• Bessonov, S., Anokhina, M., Will, C. L., Urlaub, H. & Lührmann, R. Isolation of an active step I spliceosome and composition of its RNP core. Nature 452, 846–850 (2008)
  ADS CAS PubMed Google Scholar
• Alexandrov, A., Colognori, D., Shu, M.-D. & Steitz, J. A. Human spliceosomal protein CWC22 plays a role in coupling splicing to exon junction complex deposition and nonsense-mediated decay. Proc. Natl Acad. Sci. USA 109, 21313–21318 (2012)
  ADS CAS PubMed PubMed Central Google Scholar
• Steckelberg, A.-L., Boehm, V., Gromadzka, A. M. & Gehring, N. H. CWC22 connects pre-mRNA splicing and exon junction complex assembly. Cell Reports 2, 454–461 (2012)
  CAS PubMed Google Scholar
• Barbosa, I. et al. Human CWC22 escorts the helicase eIF4AIII to spliceosomes and promotes exon junction complex assembly. Nat. Struct. Mol. Biol. 19, 983–990 (2012)
  CAS PubMed Google Scholar
• Bono, F., Ebert, J., Lorentzen, E. & Conti, E. The crystal structure of the exon junction complex reveals how it maintains a stable grip on mRNA. Cell 126, 713–725 (2006)
  CAS PubMed Google Scholar
• Andersen, C. B. F. et al. Structure of the exon junction core complex with a trapped DEAD-box ATPase bound to RNA. Science 313, 1968–1972 (2006)
  ADS CAS PubMed Google Scholar
• Davis, I. W., Murray, L. W., Richardson, J. S. & Richardson, D. C. MOLPROBITY: structure validation and all-atom contact analysis for nucleic acids and their complexes. Nucleic Acids Res. 32, W615–W619 (2004)
  CAS PubMed PubMed Central Google Scholar