DHX15-independent roles for TFIP11 in U6 snRNA modification, U4/U6.U5 tri-snRNP assembly and pre-mRNA splicing fidelity.
Cell Nucleolus
/ metabolism
Cell Survival
Coiled Bodies
/ metabolism
HeLa Cells
Humans
Methylation
Mitosis
Nuclear Proteins
/ metabolism
Nuclear Speckles
/ metabolism
Protein Binding
Protein Stability
RNA Precursors
/ metabolism
RNA Splicing
/ physiology
RNA Splicing Factors
/ genetics
RNA, Small Nuclear
/ metabolism
RNA, Small Nucleolar
/ metabolism
Ribonucleoprotein, U4-U6 Small Nuclear
/ metabolism
Ribonucleoprotein, U5 Small Nuclear
/ metabolism
Spliceosomes
/ metabolism
Journal
Nature communications
ISSN: 2041-1723
Titre abrégé: Nat Commun
Pays: England
ID NLM: 101528555
Informations de publication
Date de publication:
17 11 2021
17 11 2021
Historique:
received:
04
12
2020
accepted:
22
10
2021
entrez:
18
11
2021
pubmed:
19
11
2021
medline:
16
12
2021
Statut:
epublish
Résumé
The U6 snRNA, the core catalytic component of the spliceosome, is extensively modified post-transcriptionally, with 2'-O-methylation being most common. However, how U6 2'-O-methylation is regulated remains largely unknown. Here we report that TFIP11, the human homolog of the yeast spliceosome disassembly factor Ntr1, localizes to nucleoli and Cajal Bodies and is essential for the 2'-O-methylation of U6. Mechanistically, we demonstrate that TFIP11 knockdown reduces the association of U6 snRNA with fibrillarin and associated snoRNAs, therefore altering U6 2'-O-methylation. We show U6 snRNA hypomethylation is associated with changes in assembly of the U4/U6.U5 tri-snRNP leading to defects in spliceosome assembly and alterations in splicing fidelity. Strikingly, this function of TFIP11 is independent of the RNA helicase DHX15, its known partner in yeast. In sum, our study demonstrates an unrecognized function for TFIP11 in U6 snRNP modification and U4/U6.U5 tri-snRNP assembly, identifying TFIP11 as a critical spliceosome assembly regulator.
Identifiants
pubmed: 34789764
doi: 10.1038/s41467-021-26932-2
pii: 10.1038/s41467-021-26932-2
pmc: PMC8599867
doi:
Substances chimiques
Nuclear Proteins
0
RNA Precursors
0
RNA Splicing Factors
0
RNA, Small Nuclear
0
RNA, Small Nucleolar
0
Ribonucleoprotein, U4-U6 Small Nuclear
0
Ribonucleoprotein, U5 Small Nuclear
0
TFIP11 protein, human
0
U6 small nuclear RNA
0
p80-coilin
136882-81-0
Types de publication
Journal Article
Research Support, Non-U.S. Gov't
Langues
eng
Sous-ensembles de citation
IM
Pagination
6648Informations de copyright
© 2021. The Author(s).
Références
Wang, E. & Aifantis, I. RNA splicing and cancer. Trends Cancer 6, 631–644 (2020).
pubmed: 32434734
doi: 10.1016/j.trecan.2020.04.011
Will, C. L. & Lührmann, R. Spliceosome structure and function. Cold Spring Harb. Perspect. Biol. 3, a003707 (2011).
Paushkin, S., Gubitz, A. K., Massenet, S. & Dreyfuss, G. The SMN complex, an assemblyosome of ribonucleoproteins. Curr. Opin. Cell Biol. 14, 305–312 (2002).
pubmed: 12067652
doi: 10.1016/S0955-0674(02)00332-0
Darzacq, X. et al. Cajal body-specific small nuclear RNAs: a novel class of 2’-O-methylation and pseudouridylation guide RNAs. EMBO J. 21, 2746–2756 (2002).
pubmed: 12032087
pmcid: 126017
doi: 10.1093/emboj/21.11.2746
Didychuk, A. L., Butcher, S. E. & Brow, D. A. The life of U6 small nuclear RNA, from cradle to grave. RNA 24, 437–460 (2018).
pubmed: 29367453
pmcid: 5855946
doi: 10.1261/rna.065136.117
Dönmez, G., Hartmuth, K. & Lührmann, R. Modified nucleotides at the 5’ end of human U2 snRNA are required for spliceosomal E-complex formation. RNA 10, 1925–1933 (2004).
pubmed: 15525712
pmcid: 1370681
doi: 10.1261/rna.7186504
Karijolich, J. & Yu, Y.-T. Spliceosomal snRNA modifications and their function. RNA Biol. 7, 192–204 (2010).
pubmed: 20215871
doi: 10.4161/rna.7.2.11207
Massenet, S., Bertrand, E. & Verheggen, C. Assembly and trafficking of box C/D and H/ACA snoRNPs. RNA Biol. 14, 680–692 (2017).
pubmed: 27715451
doi: 10.1080/15476286.2016.1243646
Liu, S., Rauhut, R., Vornlocher, H.-P. & Lührmann, R. The network of protein-protein interactions within the human U4/U6.U5 tri-snRNP. RNA 12, 1418–1430 (2006).
pubmed: 16723661
pmcid: 1484429
doi: 10.1261/rna.55406
Schaffert, N., Hossbach, M., Heintzmann, R., Achsel, T. & Lührmann, R. RNAi knockdown of hPrp31 leads to an accumulation of U4/U6 di-snRNPs in Cajal bodies. EMBO J. 23, 3000–3009 (2004).
pubmed: 15257298
pmcid: 514917
doi: 10.1038/sj.emboj.7600296
Liu, S. et al. A composite double-/single-stranded RNA-binding region in protein Prp3 supports tri-snRNP stability and splicing. Elife 4, e07320 (2015).
Tanaka, N., Aronova, A. & Schwer, B. Ntr1 activates the Prp43 helicase to trigger release of lariat-intron from the spliceosome. Genes Dev. 21, 2312–2325 (2007).
pubmed: 17875666
pmcid: 1973145
doi: 10.1101/gad.1580507
Fourmann, J.-B., Tauchert, M. J., Ficner, R., Fabrizio, P. & Lührmann, R. Regulation of Prp43-mediated disassembly of spliceosomes by its cofactors Ntr1 and Ntr2. Nucleic Acids Res. 45, 4068–4080 (2017).
pubmed: 27923990
doi: 10.1093/nar/gkw1225
Christian, H., Hofele, R. V., Urlaub, H. & Ficner, R. Insights into the activation of the helicase Prp43 by biochemical studies and structural mass spectrometry. Nucleic Acids Res. 42, 1162–1179 (2014).
pubmed: 24165877
doi: 10.1093/nar/gkt985
Yoshimoto, R., Kataoka, N., Okawa, K. & Ohno, M. Isolation and characterization of post-splicing lariat–intron complexes. Nucleic Acids Res. 37, 891–902 (2009).
pubmed: 19103666
doi: 10.1093/nar/gkn1002
Memet, I., Doebele, C., Sloan, K. E. & Bohnsack, M. T. The G-patch protein NF-κB-repressing factor mediates the recruitment of the exonuclease XRN2 and activation of the RNA helicase DHX15 in human ribosome biogenesis. Nucleic Acids Res. 45, 5359–5374 (2017).
pubmed: 28115624
pmcid: 5435916
Fakan, S. Perichromatin fibrils are in situ forms of nascent transcripts. Trends Cell Biol. 4, 86–90 (1994).
pubmed: 14731598
doi: 10.1016/0962-8924(94)90180-5
Strzelecka, M. et al. Coilin-dependent snRNP assembly is essential for zebrafish embryogenesis. Nat. Struct. Mol. Biol. 17, 403–409 (2010).
pubmed: 20357773
doi: 10.1038/nsmb.1783
Uversky, V. N. Intrinsically disordered proteins and their “mysterious” (meta)physics. Front. Phys. 7, 10 (2019).
doi: 10.3389/fphy.2019.00010
Nygaard, M., Kragelund, B. B., Papaleo, E. & Lindorff-Larsen, K. An efficient method for estimating the hydrodynamic radius of disordered protein conformations. Biophys. J. 113, 550–557 (2017).
pubmed: 28793210
pmcid: 5550300
doi: 10.1016/j.bpj.2017.06.042
Wootton, J. C. Non-globular domains in protein sequences: automated segmentation using complexity measures. Comput. Chem. 18, 269–285 (1994).
pubmed: 7952898
doi: 10.1016/0097-8485(94)85023-2
Frege, T. & Uversky, V. N. Intrinsically disordered proteins in the nucleus of human cells. Biochem. Biophys. Rep. 1, 33 (2015).
pubmed: 29124132
pmcid: 5668563
Lemm, I. et al. Ongoing U snRNP biogenesis is required for the integrity of Cajal bodies. Mol. Biol. Cell 17, 3221–3231 (2006).
pubmed: 16687569
pmcid: 1483051
doi: 10.1091/mbc.e06-03-0247
Novotný, I. et al. SART3-dependent accumulation of incomplete spliceosomal snRNPs in cajal bodies. Cell Rep. 10, 429–440 (2015).
pubmed: 25600876
doi: 10.1016/j.celrep.2014.12.030
Bohmann, K., Ferreira, J. A. & Lamond, A. I. Mutational analysis of p80 coilin indicates a functional interaction between coiled bodies and the nucleolus. J. Cell Biol. 131, 817–831 (1995).
pubmed: 7490287
doi: 10.1083/jcb.131.4.817
Le Tonquèze, O., Gschloessl, B., Legagneux, V., Paillard, L. & Audic, Y. Identification of CELF1 RNA targets by CLIP-seq in human HeLa cells. Genomics Data 8, 97–103 (2016).
pubmed: 27222809
pmcid: 4872370
doi: 10.1016/j.gdata.2016.04.009
Machyna, M. et al. The coilin interactome identifies hundreds of small noncoding RNAs that traffic through cajal bodies. Mol. Cell 56, 389–399 (2014).
pubmed: 25514182
doi: 10.1016/j.molcel.2014.10.004
Wang, Z. et al. iCLIP predicts the dual splicing effects of TIA-RNA interactions. PLoS Biol. 8, e1000530 (2010).
pubmed: 21048981
pmcid: 2964331
doi: 10.1371/journal.pbio.1000530
Zarnack, K. et al. Direct competition between hnRNP C and U2AF65 protects the transcriptome from the exonization of Alu elements. Cell 152, 453–466 (2013).
pubmed: 23374342
pmcid: 3629564
doi: 10.1016/j.cell.2012.12.023
Hutten, S., Chachami, G., Winter, U., Melchior, F. & Lamond, A. I. A role for the Cajal-body-associated SUMO isopeptidase USPL1 in snRNA transcription mediated by RNA polymerase II. J. Cell Sci. 127, 1065–1078 (2014).
pubmed: 24413172
pmcid: 3937775
Makarova, O. V., Makarov, E. M. & Lührmann, R. The 65 and 110 kDa SR-related proteins of the U4/U6.U5 tri-snRNP are essential for the assembly of mature spliceosomes. EMBO J. 20, 2553–2563 (2001).
pubmed: 11350945
pmcid: 125249
doi: 10.1093/emboj/20.10.2553
Sander, B. et al. Organization of core spliceosomal components U5 snRNA Loop I and U4/U6 Di-snRNP within U4/U6.U5 Tri-snRNP as revealed by electron cryomicroscopy. Mol. Cell 24, 267–278.
Achsel, T., Ahrens, K., Brahms, H., Teigelkamp, S. & Lührmann, R. The human U5-220kD protein (hPrp8) forms a stable RNA-free complex with several U5-specific proteins, including an RNA unwindase, a homologue of ribosomal elongation factor EF-2, and a novel WD-40 protein. Mol. Cell. Biol. 18, 6756–6766 (1998).
pubmed: 9774689
pmcid: 109259
doi: 10.1128/MCB.18.11.6756
Anthony, J. G., Weidenhammer, E. M. & Woolford, J. L. The yeast Prp3 protein is a U4/U6 snRNP protein necessary for integrity of the U4/U6 snRNP and the U4/U6.U5 tri-snRNP. RNA 3, 1143–1152 (1997).
pubmed: 9326489
pmcid: 1369556
Song, E. J. et al. The Prp19 complex and the Usp4Sart3 deubiquitinating enzyme control reversible ubiquitination at the spliceosome. Genes Dev. 24, 1434–1447 (2010).
pubmed: 20595234
pmcid: 2895201
doi: 10.1101/gad.1925010
Pozzi, B. et al. SUMO conjugation to spliceosomal proteins is required for efficient pre-mRNA splicing. Nucleic Acids Res. 45, 6729–6745 (2017).
pubmed: 28379520
pmcid: 5499870
doi: 10.1093/nar/gkx213
Birkedal, U. et al. Profiling of ribose methylations in RNA by high-throughput sequencing. Angew. Chem. Int. Ed. Engl. 54, 451–455 (2015).
Marchand, V., Blanloeil-Oillo, F., Helm, M. & Motorin, Y. Illumina-based RiboMethSeq approach for mapping of 2’-O-Me residues in RNA. Nucleic Acids Res. 44, e135 (2016).
pubmed: 27302133
pmcid: 5027498
doi: 10.1093/nar/gkw547
Nachmani, D. et al. Germline NPM1 mutations lead to altered rRNA 2’-O-methylation and cause dyskeratosis congenita. Nat. Genet. 51, 1518–1529 (2019).
pubmed: 31570891
pmcid: 6858547
doi: 10.1038/s41588-019-0502-z
Uhlmann, F. Chromosome cohesion and segregation in mitosis and meiosis. Curr. Opin. Cell Biol. 13, 754–761 (2001).
pubmed: 11698193
doi: 10.1016/S0955-0674(00)00279-9
Valcárcel, J. & Malumbres, M. Splicing together sister chromatids. EMBO J. 33, 2601–2603 (2014).
pubmed: 25266476
pmcid: 4282569
doi: 10.15252/embj.201489988
Wang, X. et al. LARP7-mediated U6 snRNA modification ensures splicing fidelity and spermatogenesis in mice. Mol. Cell 77, 999–1013 (2020). e6.
pubmed: 32017896
doi: 10.1016/j.molcel.2020.01.002
Hasler, D. et al. The Alazami syndrome-associated protein LARP7 guides U6 small nuclear RNA modification and contributes to splicing robustness. Mol. Cell 77, 1014–1031.e13 (2020).
D’Souza, M. N. et al. FMRP interacts with C/D Box snoRNA in the nucleus and regulates ribosomal RNA methylation. iScience 9, 399–411 (2018).
pubmed: 30469012
pmcid: 6249352
doi: 10.1016/j.isci.2018.11.007
Nottrott, S., Urlaub, H. & Lührmann, R. Hierarchical, clustered protein interactions with U4/U6 snRNA: a biochemical role for U4/U6 proteins. EMBO J. 21, 5527 (2002).
pubmed: 12374753
pmcid: 129076
doi: 10.1093/emboj/cdf544
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).
pubmed: 9705931
doi: 10.1016/S0960-9822(07)00345-4
Rodgers, M. L., Didychuk, A. L., Butcher, S. E., Brow, D. A. & Hoskins, A. A. A multi-step model for facilitated unwinding of the yeast U4/U6 RNA duplex. Nucleic Acids Res. 44, 10912–10928 (2016).
pubmed: 27484481
pmcid: 5159527
doi: 10.1093/nar/gkw686
Sidarovich, A. et al. Identification of a small molecule inhibitor that stalls splicing at an early step of spliceosome activation. Elife 6, e23533 (2017).
Deckert, J. et al. Protein composition and electron microscopy structure of affinity-purified human spliceosomal B complexes isolated under physiological conditions. Mol. Cell. Biol. 26, 5528 (2006).
pubmed: 16809785
pmcid: 1592722
doi: 10.1128/MCB.00582-06
Dvinge, H., Guenthoer, J., Porter, P. L. & Bradley, R. K. RNA components of the spliceosome regulate tissue- and cancer-specific alternative splicing. Genome Res. 29, 1591–1604.
Niu, Z., Jin, W., Zhang, L. & Li, X. Tumor suppressor RBM5 directly interacts with the DExD/H-box protein DHX15 and stimulates its helicase activity. FEBS Lett. 586, 977–983 (2012).
pubmed: 22569250
doi: 10.1016/j.febslet.2012.02.052
Lin, M.-L. et al. Involvement of G-patch domain containing 2 overexpression in breast carcinogenesis. Cancer Sci. 100, 1443–1450 (2009).
pubmed: 19432882
doi: 10.1111/j.1349-7006.2009.01185.x
Chen, Y.-L. et al. The telomerase inhibitor Gno1p/PINX1 activates the helicase Prp43p during ribosome biogenesis. Nucleic Acids Res. 42, 7330–7345 (2014).
pubmed: 24823796
pmcid: 4066782
doi: 10.1093/nar/gku357
Bohnsack, M. T. et al. Prp43 bound at different sites on the pre-rRNA performs distinct functions in ribosome synthesis. Mol. Cell 36, 583–592 (2009).
pubmed: 19941819
pmcid: 2806949
doi: 10.1016/j.molcel.2009.09.039
Uchiyama, S. & Fukui, K. Condensin in chromatid cohesion and segregation. Cytogenet. Genome Res. 147, 212–216 (2015).
pubmed: 26998746
doi: 10.1159/000444868
Zhang, L. et al. Conserved eukaryotic kinase CK2 chaperone intrinsically disordered protein interactions. Appl. Environ. Microbiol. 86, e02191–19 (2020).
Tannukit, S. et al. Identification of a novel nuclear localization signal and speckle-targeting sequence of tuftelin-interacting protein 11, a splicing factor involved in spliceosome disassembly. Biochem. Biophys. Res. Commun. 390, 1044–1050 (2009).
pubmed: 19857462
pmcid: 2787706
doi: 10.1016/j.bbrc.2009.10.111
Santofimia-Castaño, P. et al. Targeting intrinsically disordered proteins involved in cancer. Cell. Mol. Life Sci. 77, 1695–1707 (2020).
pubmed: 31667555
doi: 10.1007/s00018-019-03347-3
Huppertz, I. et al. iCLIP: protein-RNA interactions at nucleotide resolution. Methods 65, 274–287 (2014).
pubmed: 24184352
pmcid: 3988997
doi: 10.1016/j.ymeth.2013.10.011
de Araujo Oliveira, J. V. et al. SnoReport 2.0: new features and a refined Support Vector Machine to improve snoRNA identification. BMC Bioinformatics 17, 73–86 (2016).
doi: 10.1186/s12859-016-1345-6
Signal, B., Gloss, B. S., Dinger, M. E. & Mercer, T. R. Machine learning annotation of human branchpoints. Bioinformatics 34, 920–927 (2018).
pubmed: 29092009
doi: 10.1093/bioinformatics/btx688
Huang, D. W. et al. The DAVID Gene Functional Classification Tool: a novel biological module-centric algorithm to functionally analyze large gene lists. Genome Biol. 8, R183 (2007).
pubmed: 17784955
pmcid: 2375021
doi: 10.1186/gb-2007-8-9-r183
Lestrade, L. & Weber, M. J. snoRNA-LBME-db, a comprehensive database of human H/ACA and C/D box snoRNAs. Nucleic Acids Res. 34, D158–D162 (2006).
doi: 10.1093/nar/gkj002
Kyte, J. & Doolittle, R. F. A simple method for displaying the hydropathic character of a protein. J. Mol. Biol. 157, 105–132 (1982).
Yang, J. et al. The I-TASSER Suite: protein structure and function prediction. Nat. Methods 12, 7–8 (2015).
pubmed: 25549265
pmcid: 4428668
doi: 10.1038/nmeth.3213
Tomasso, M. E., Tarver, M. J., Devarajan, D. & Whitten, S. T. Hydrodynamic radii of intrinsically disordered proteins determined from experimental polyproline II propensities. PLoS Comput. Biol. 12, e1004686 (2016).
pubmed: 26727467
pmcid: 4699819
doi: 10.1371/journal.pcbi.1004686