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The human cap-binding complex is functionally connected to the nuclear RNA exosome


Nuclear processing and quality control of eukaryotic RNA is mediated by the RNA exosome, which is regulated by accessory factors. However, the mechanism of exosome recruitment to its ribonucleoprotein (RNP) targets remains poorly understood. Here we report a physical link between the human exosome and the cap-binding complex (CBC). The CBC associates with the ARS2 protein to form CBC–ARS2 (CBCA) and then further connects, together with the ZC3H18 protein, to the nuclear exosome targeting (NEXT) complex, thus forming CBC–NEXT (CBCN). RNA immunoprecipitation using CBCN factors as well as the analysis of combinatorial depletion of CBCN and exosome components underscore the functional relevance of CBC-exosome bridging at the level of target RNA. Specifically, CBCA suppresses read-through products of several RNA families by promoting their transcriptional termination. We suggest that the RNP 5′ cap links transcription termination to exosomal RNA degradation through CBCN.

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Figure 1: AC of NEXT-complex components hMTR4 and RBM7 reveals stoichiometrically interacting ZC3H18 and CBCA complex.
Figure 2: AC of CBCA complex components CBP80, CBP20 and ARS2 reveals stoichiometric CBCN complex.
Figure 3: AC of ZC3H18 identifies stoichiometric CBCN complex.
Figure 4: CBCN components share common RNA targets.
Figure 5: Hyperaccumulation of PROMPTs upon co-depletion of CBCA with components of NEXT or RNA-exosome complexes.
Figure 6: CBCA depletion causes inefficient transcription termination and exosome-dependent removal of read-through transcripts.
Figure 7: CBCA depletion displays a genome-wide termination defect of PROMPT transcription.

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  1. Chlebowski, A., Lubas, M., Jensen, T.H. & Dziembowski, A. RNA decay machines: the exosome. Biochim. Biophys. Acta 1829, 552–560 (2013).

    Article  CAS  Google Scholar 

  2. Houseley, J. & Tollervey, D. The many pathways of RNA degradation. Cell 136, 763–776 (2009).

    Article  CAS  Google Scholar 

  3. Staals, R.H. et al. Dis3-like 1: a novel exoribonuclease associated with the human exosome. EMBO J. 29, 2358–2367 (2010).

    Article  CAS  Google Scholar 

  4. Tomecki, R. et al. The human core exosome interacts with differentially localized processive RNases: hDIS3 and hDIS3L. EMBO J. 29, 2342–2357 (2010).

    Article  CAS  Google Scholar 

  5. Lykke-Andersen, S., Brodersen, D.E. & Jensen, T.H. Origins and activities of the eukaryotic exosome. J. Cell Sci. 122, 1487–1494 (2009).

    Article  CAS  Google Scholar 

  6. Lubas, M. et al. Interaction profiling identifies the human nuclear exosome targeting complex. Mol. Cell 43, 624–637 (2011).

    Article  CAS  Google Scholar 

  7. LaCava, J. et al. RNA degradation by the exosome is promoted by a nuclear polyadenylation complex. Cell 121, 713–724 (2005).

    Article  CAS  Google Scholar 

  8. Wyers, F. et al. Cryptic pol II transcripts are degraded by a nuclear quality control pathway involving a new poly(A) polymerase. Cell 121, 725–737 (2005).

    Article  CAS  Google Scholar 

  9. Vanácová, S. et al. A new yeast poly(A) polymerase complex involved in RNA quality control. PLoS Biol. 3, e189 (2005).

    Article  Google Scholar 

  10. Preker, P. et al. RNA exosome depletion reveals transcription upstream of active human promoters. Science 322, 1851–1854 (2008).

    Article  CAS  Google Scholar 

  11. Arigo, J.T., Eyler, D.E., Carroll, K.L. & Corden, J.L. Termination of cryptic unstable transcripts is directed by yeast RNA-binding proteins Nrd1 and Nab3. Mol. Cell 23, 841–851 (2006).

    Article  CAS  Google Scholar 

  12. Thiebaut, M., Kisseleva-Romanova, E., Rougemaille, M., Boulay, J. & Libri, D. Transcription termination and nuclear degradation of cryptic unstable transcripts: a role for the nrd1-nab3 pathway in genome surveillance. Mol. Cell 23, 853–864 (2006).

    Article  CAS  Google Scholar 

  13. Vasiljeva, L. & Buratowski, S. Nrd1 interacts with the nuclear exosome for 3′ processing of RNA polymerase II transcripts. Mol. Cell 21, 239–248 (2006).

    Article  CAS  Google Scholar 

  14. Steinmetz, E.J., Conrad, N.K., Brow, D.A. & Corden, J.L. RNA-binding protein Nrd1 directs poly(A)-independent 3′-end formation of RNA polymerase II transcripts. Nature 413, 327–331 (2001).

    Article  CAS  Google Scholar 

  15. Gruber, J.J. et al. Ars2 links the nuclear cap-binding complex to RNA interference and cell proliferation. Cell 138, 328–339 (2009).

    Article  CAS  Google Scholar 

  16. Izaurralde, E. et al. A nuclear cap binding protein complex involved in pre-mRNA splicing. Cell 78, 657–668 (1994).

    Article  CAS  Google Scholar 

  17. Rasmussen, E.B. & Lis, J.T. In vivo transcriptional pausing and cap formation on three Drosophila heat shock genes. Proc. Natl. Acad. Sci. USA 90, 7923–7927 (1993).

    Article  CAS  Google Scholar 

  18. Görnemann, J., Kotovic, K.M., Hujer, K. & Neugebauer, K.M. Cotranscriptional spliceosome assembly occurs in a stepwise fashion and requires the cap binding complex. Mol. Cell 19, 53–63 (2005).

    Article  Google Scholar 

  19. Flaherty, S.M., Fortes, P., Izaurralde, E., Mattaj, I.W. & Gilmartin, G.M. Participation of the nuclear cap binding complex in pre-mRNA 3′ processing. Proc. Natl. Acad. Sci. USA 94, 11893–11898 (1997).

    Article  CAS  Google Scholar 

  20. Hosoda, N., Kim, Y.K., Lejeune, F. & Maquat, L.E. CBP80 promotes interaction of Upf1 with Upf2 during nonsense-mediated mRNA decay in mammalian cells. Nat. Struct. Mol. Biol. 12, 893–901 (2005).

    Article  CAS  Google Scholar 

  21. Izaurralde, E. et al. A cap-binding protein complex mediating U snRNA export. Nature 376, 709–712 (1995).

    Article  CAS  Google Scholar 

  22. Cheng, H. et al. Human mRNA export machinery recruited to the 5′ end of mRNA. Cell 127, 1389–1400 (2006).

    Article  CAS  Google Scholar 

  23. Ohno, M., Segref, A., Bachi, A., Wilm, M. & Mattaj, I.W. PHAX, a mediator of U snRNA nuclear export whose activity is regulated by phosphorylation. Cell 101, 187–198 (2000).

    Article  CAS  Google Scholar 

  24. Kataoka, N., Ohno, M., Moda, I. & Shimura, Y. Identification of the factors that interact with NCBP, an 80 kDa nuclear cap binding protein. Nucleic Acids Res. 23, 3638–3641 (1995).

    Article  CAS  Google Scholar 

  25. Boulon, S. et al. PHAX and CRM1 are required sequentially to transport U3 snoRNA to nucleoli. Mol. Cell 16, 777–787 (2004).

    Article  CAS  Google Scholar 

  26. Balatsos, N.A., Nilsson, P., Mazza, C., Cusack, S. & Virtanen, A. Inhibition of mRNA deadenylation by the nuclear cap binding complex (CBC). J. Biol. Chem. 281, 4517–4522 (2006).

    Article  CAS  Google Scholar 

  27. Domanski, M. et al. Improved methodology for the affinity isolation of human protein complexes expressed at near endogenous levels. Biotechniques 0, 1–6 (2012).

    PubMed  PubMed Central  Google Scholar 

  28. Poser, I. et al. BAC TransgeneOmics: a high-throughput method for exploration of protein function in mammals. Nat. Methods 5, 409–415 (2008).

    Article  CAS  Google Scholar 

  29. Hubner, N.C. & Mann, M. Extracting gene function from protein-protein interactions using Quantitative BAC InteraCtomics (QUBIC). Methods 53, 453–459 (2011).

    Article  CAS  Google Scholar 

  30. Kiriyama, M., Kobayashi, Y., Saito, M., Ishikawa, F. & Yonehara, S. Interaction of FLASH with arsenite resistance protein 2 is involved in cell cycle progression at S phase. Mol. Cell Biol. 29, 4729–4741 (2009).

    Article  CAS  Google Scholar 

  31. Preker, P. et al. PROMoter uPstream Transcripts share characteristics with mRNAs and are produced upstream of all three major types of mammalian promoters. Nucleic Acids Res. 39, 7179–7193 (2011).

    Article  CAS  Google Scholar 

  32. Ntini, E. et al. Polyadenylation site-induced decay of upstream transcripts enforces promoter directionality. Nat. Struct. Mol. Biol. 20, 923–928 (2013).

    Article  CAS  Google Scholar 

  33. Hallais, M. et al. CBC-ARS2 stimulate 3′-end maturation of multiple RNA families and favor cap-proximal processing. Nat. Struct. Mol. Biol. 10.1038/nsmb.2720 (24 November 2013).

  34. Andrulis, E.D. et al. The RNA processing exosome is linked to elongating RNA polymerase II in Drosophila. Nature 420, 837–841 (2002).

    Article  CAS  Google Scholar 

  35. Hessle, V. et al. The exosome associates cotranscriptionally with the nascent pre-mRNP through interactions with heterogeneous nuclear ribonucleoproteins. Mol. Biol. Cell 20, 3459–3470 (2009).

    Article  CAS  Google Scholar 

  36. Hieronymus, H., Yu, M.C. & Silver, P.A. Genome-wide mRNA surveillance is coupled to mRNA export. Genes Dev. 18, 2652–2662 (2004).

    Article  CAS  Google Scholar 

  37. Das, B., Butler, J.S. & Sherman, F. Degradation of normal mRNA in the nucleus of Saccharomyces cerevisiae. Mol. Cell Biol. 23, 5502–5515 (2003).

    Article  CAS  Google Scholar 

  38. Kuai, L., Das, B. & Sherman, F. A nuclear degradation pathway controls the abundance of normal mRNAs in Saccharomyces cerevisiae. Proc. Natl. Acad. Sci. USA 102, 13962–13967 (2005).

    Article  CAS  Google Scholar 

  39. Visa, N., Izaurralde, E., Ferreira, J., Daneholt, B. & Mattaj, I.W. A nuclear cap-binding complex binds Balbiani ring pre-mRNA cotranscriptionally and accompanies the ribonucleoprotein particle during nuclear export. J. Cell Biol. 133, 5–14 (1996).

    Article  CAS  Google Scholar 

  40. Glover-Cutter, K., Kim, S., Espinosa, J. & Bentley, D.L. RNA polymerase II pauses and associates with pre-mRNA processing factors at both ends of genes. Nat. Struct. Mol. Biol. 15, 71–78 (2008).

    Article  CAS  Google Scholar 

  41. Listerman, I., Sapra, A.K. & Neugebauer, K.M. Cotranscriptional coupling of splicing factor recruitment and precursor messenger RNA splicing in mammalian cells. Nat. Struct. Mol. Biol. 13, 815–822 (2006).

    Article  CAS  Google Scholar 

  42. Andreu-Agullo, C., Maurin, T., Thompson, C.B. & Lai, E.C. Ars2 maintains neural stem-cell identity through direct transcriptional activation of Sox2. Nature 481, 195–198 (2012).

    Article  CAS  Google Scholar 

  43. Shi, Y. et al. Molecular architecture of the human pre-mRNA 3′ processing complex. Mol. Cell 33, 365–376 (2009).

    Article  CAS  Google Scholar 

  44. Narita, T. et al. NELF interacts with CBC and participates in 3′ end processing of replication-dependent histone mRNAs. Mol. Cell 26, 349–365 (2007).

    Article  CAS  Google Scholar 

  45. Gruber, J.J. et al. Ars2 promotes proper replication-dependent histone mRNA 3′ end formation. Mol. Cell 45, 87–98 (2012).

    Article  CAS  Google Scholar 

  46. Yang, X.C., Burch, B.D., Yan, Y., Marzluff, W.F. & Dominski, Z. FLASH, a proapoptotic protein involved in activation of caspase-8, is essential for 3′ end processing of histone pre-mRNAs. Mol. Cell 36, 267–278 (2009).

    Article  CAS  Google Scholar 

  47. Lenasi, T., Peterlin, B.M. & Barboric, M. Cap-binding protein complex links pre-mRNA capping to transcription elongation and alternative splicing through positive transcription elongation factor b (P-TEFb). J. Biol. Chem. 286, 22758–22768 (2011).

    Article  CAS  Google Scholar 

  48. Das, R. et al. SR proteins function in coupling RNAP II transcription to pre-mRNA splicing. Mol. Cell 26, 867–881 (2007).

    Article  CAS  Google Scholar 

  49. Grzechnik, P. & Kufel, J. Polyadenylation linked to transcription termination directs the processing of snoRNA precursors in yeast. Mol. Cell 32, 247–258 (2008).

    Article  CAS  Google Scholar 

  50. Rondón, A.G., Mischo, H.E., Kawauchi, J. & Proudfoot, N.J. Fail-safe transcriptional termination for protein-coding genes in S. cerevisiae. Mol. Cell 36, 88–98 (2009).

    Article  Google Scholar 

  51. Almada, A.E., Wu, X., Kriz, A.J., Burge, C.B. & Sharp, P.A. Promoter directionality is controlled by U1 snRNP and polyadenylation signals. Nature 499, 360–363 (2013).

    Article  CAS  Google Scholar 

  52. Gudipati, R.K., Villa, T., Boulay, J. & Libri, D. Phosphorylation of the RNA polymerase II C-terminal domain dictates transcription termination choice. Nat. Struct. Mol. Biol. 15, 786–794 (2008).

    Article  CAS  Google Scholar 

  53. Vasiljeva, L., Kim, M., Mutschler, H., Buratowski, S. & Meinhart, A. The Nrd1-Nab3-Sen1 termination complex interacts with the Ser5-phosphorylated RNA polymerase II C-terminal domain. Nat. Struct. Mol. Biol. 15, 795–804 (2008).

    Article  CAS  Google Scholar 

  54. Wiśniewski, J.R., Zougman, A., Nagaraj, N. & Mann, M. Universal sample preparation method for proteome analysis. Nat. Methods 6, 359–362 (2009).

    Article  Google Scholar 

  55. Dobin, A. et al. STAR: ultrafast universal RNA-seq aligner. Bioinformatics 29, 15–21 (2013).

    Article  CAS  Google Scholar 

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We thank M. Schmid, S. Lykke-Andersen, M. Lubas, A. Dziembowski and D. Libri for critical comments to the manuscript. Special thanks go to E. Marchal for excellent technical assistance. E. Izaurralde (Max Planck Institute for Developmental Biology) and J. Lykke-Andersen (Division of Biological Sciences at University of California, San Diego) are acknowledged for sharing reagents. This work was supported by the Danish National Research Foundation (grant DNRF58), the Danish Cancer Society and the Lundbeck and Novo Nordisk Foundations (T.H.J.); the US National Institutes of Health (grant U54 GM103511, to M.P.R.), the l'ARC and La Ligue Contre Le Cancer (E.B.), the Foundation for Strategic Research (grant FFL09-0130, to R.S.) and the European Community's Seventh Framework Programme (FP7/2007-2013) under grant no. 241548 (MitoSys, to A.H.).

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Authors and Affiliations



M.D. and J.B. performed ACMS experiments and analyzed the data together with J.S.A., J.L. and M.P.R.; I.P. and A.H. contributed tagged cell lines. C.V., M.H. and E.B. performed and analyzed RIP experiments. P.R.A., M.S.K. and A.S. performed experimental RNA analyses. P.R.A. and E.N. performed and analyzed ChIP experiments. P.R.A., H.S. and R.S. analyzed RNA-seq data. P.R.A., M.D., M.S.K. and T.H.J. wrote the manuscript.

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Correspondence to Torben Heick Jensen.

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The authors declare no competing financial interests.

Integrated supplementary information

Supplementary Figure 1 Validation of near-endogenous expression levels of tagged proteins.

Western blotting analyses of tagged proteins used for AC/MS analyses as well as their endogenous counterparts. Membranes were probed with antibodies targeting the indicated factors. Tagged versions of CBP20, CBP80 and ZC3H18 were expressed at endogenous levels, while tagged versions of ARS2 and hMTR4 were expressed at lower levels than their endogenous versions. RBM7-LAP was ~2-fold overexpressed, which is unlikely to have caused any artificial interactions due to the high level of congruency between AC/MS analyses of RBM7-LAP, hMTR4-LAP and ZCCHC8-FLAG6

Supplementary Figure 2 Co-depletion of major 5'-3' exonucleolytic enzymes does not affect PROMPT levels.

(a) Western blotting analysis of whole cell extracts showing protein depletion upon the indicated siRNA administrations. Membranes were probed with the indicated antibodies. Anti-actin antibody was used as a loading control. (b) XRN1 + XRN2-depletion does not increase PROMPT levels of exosome-depleted cells. Levels of the indicated PROMPTs were measured and displayed as in Fig. 5b.

Supplementary Figure 3 Effect of depletion of CBCA components on transcription termination of the U1- and U8-encoding genes.

(a) Transcriptional read-through analysis of the U1 gene upon the indicated factor depletions and displayed as in Fig. 6b. (b) Northern blotting analysis of total RNA purified from HeLa cells subjected to the indicated siRNA transfections (depletion levels shown in Fig. 5a). Position of the utilized 5'end radiolabeled probe to primarily detect 3'extended U8 RNA species is shown at the top. Factor depletions (Fig. 5a), data display and control were done as in Fig. 5c. (c) Transcriptional read-through analysis of the U8 gene upon the indicated factor depletions and displayed as in Fig. 6b.

Supplementary Figure 4 Mean coverage ratios around TSSs upon CBCN factor depletion.

Plots are displayed similarly to those in Figure 7. (a) PROMPT accumulation upon combinatorial depletion of hRRP40 and CBCN relative to control siRNA. (b) PROMPT accumulation upon combinatorial depletion of ZCCHC8 and CBCN relative to control siRNA. (c) PROMPT accumulation upon combinatorial depletion of ZCCHC8 and CBCN relative to ZCCHC8 depletion. Faded areas represent 95% confidence intervals for each curve.

Supplementary Figure 5 Mean coverage ratios around histone mRNA TSSs upon CBCN factor depletion.

Plots are displayed similarly to those in Fig. 7. For a-d upper panels display the region around annotated 3'ends of mature replication-dependent histone (RDH) genes, while lower panels display that of replication-independent histone (RIH) genes. (a) Mean coverage as tags per event (TPE) at histone gene TTSs. (b) Histone RNA accumulation upon single factor depletion relative to control siRNA. (c) Histone RNA accumulation upon combinatorial depletion of hRRP40 and CBCN relative to control siRNA. (d) Histone RNA accumulation upon combinatorial depletion of hRRP40 and CBCN relative to hRRP40 depletion.

Supplementary Figure 6 Uncropped gel images of northern blots in Figure 5 and Supplementary Figure 3.

(a) Uncropped image of the northern blot shown in Fig. 5c. (b) Uncropped image of the northern blot shown in Supplementary Fig. 3b.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–6, Supplementary Tables 1 and 8–13 and Supplementary Note (PDF 1893 kb)

Supplementary Table 2


Supplementary Table 3


Supplementary Table 4

CBP80–3× FLAG ACMS (XLSX 23 kb)

Supplementary Table 5

CBP20–3× FLAG ACMS (XLSX 17 kb)

Supplementary Table 6


Supplementary Table 7

ZC3H18–3× FLAG ACMS (XLSX 22 kb)

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Andersen, P., Domanski, M., Kristiansen, M. et al. The human cap-binding complex is functionally connected to the nuclear RNA exosome. Nat Struct Mol Biol 20, 1367–1376 (2013).

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