Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Article
  • Published:

Telomere-independent Rap1 is an IKK adaptor and regulates NF-κB-dependent gene expression

Nature Cell Biology volume 12, pages 758–767 (2010)Cite this article

Subjects

Abstract

We describe a genome-wide gain-of-function screen for regulators of NF-κB, and identify Rap1 (Trf2IP), as an essential modulator of NF-κB-mediated pathways. NF-κB is induced by ectopic expression of Rap1, whereas its activity is inhibited by Rap1 depletion. In addition to localizing on telomeres, mammalian Rap1 forms a complex with IKKs (IκB kinases), and is crucial for the ability of IKKs to be recruited to, and phosphorylate, the p65 subunit of NF-κB to make it transcriptionally competent. Rap1-mutant mice display defective NF-κB activation and are resistant to endotoxic shock. Furthermore, levels of Rap1 are positively regulated by NF-κB, and human breast cancers with NF-κB hyperactivity show elevated levels of cytoplasmic Rap1. Similar to inhibiting NF-κB, knockdown of Rap1 sensitizes breast cancer cells to apoptosis. These results identify the first cytoplasmic role of Rap1 and provide a mechanism through which it regulates an important signalling cascade in mammals, independent of its ability to regulate telomere function.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1: Identification of Rap1 as a regulator of NF-κB signalling.
Figure 2: Interaction of Rap1 with IKKs is required for NF-κB activation.
Figure 3: Rap1 knockdown impairs NF-κB signalling.
Figure 4: Rap1 is required for recruitment of IKK to p65 and for phosphorylation of p65 at Ser 536.
Figure 5: Rap1 levels are regulated by NF-κB signalling.
Figure 6: Rap1-mutant mice are resistant to endotoxic shock and show defective NF-κB signalling in response to LPS.
Figure 7: Rap1 staining in graded breast cancers.
Figure 8: Role of Rap1 signalling in breast cancer.

Similar content being viewed by others

References

  1. Ghosh, S. & Karin, M. Missing pieces in the NF-κB puzzle. Cell 109, S81–S96 (2002).

    Article  CAS  Google Scholar 

  2. Karin, M. & Ben-Neriah, Y. Phosphorylation meets ubiquitination: the control of NF-κB activity. Annu. Rev. Immunol. 18, 621–663 (2000).

    Article  CAS  Google Scholar 

  3. Silverman, N. & Maniatis, T. NF-κB signaling pathways in mammalian and insect innate immunity. Genes Dev. 15, 2321–2342 (2001).

    Article  CAS  Google Scholar 

  4. Sasaki, Y., Schmidt-Supprian, M., Derudder, E. & Rajewsky, K. Role of NFκB signaling in normal and malignant B cell development. Adv. Exp. Med. Biol. 596, 149–154 (2007).

    Article  Google Scholar 

  5. Pasparakis, M. Regulation of tissue homeostasis by NF-κB signalling: implications for inflammatory diseases. Nat. Rev. Immunol. 9, 778–788 (2009).

    Article  CAS  Google Scholar 

  6. Aggarwal, B. B. Nuclear factor-κB: the enemy within. Cancer Cell 6, 203–208 (2004).

    Article  CAS  Google Scholar 

  7. Tergaonkar, V., Correa, R. G., Ikawa, M. & Verma, I. M. Distinct roles of IκB proteins in regulating constitutive NF-κB activity. Nat. Cell Biol. 7, 921–923 (2005).

    Article  CAS  Google Scholar 

  8. Bates, P. W. & Miyamoto, S. Expanded nuclear roles for IκBs. Sci. STKE 2004, pe48 (2004).

    PubMed  Google Scholar 

  9. Bhoj, V. G. & Chen, Z. J. Ubiquitylation in innate and adaptive immunity. Nature 458, 430–437 (2009).

    Article  CAS  Google Scholar 

  10. Wertz, I. E & Dixit, V. M. Ubiquitin-mediated regulation of TNFR1 signaling. Cytokine Growth Factor Rev. 19, 313–324 (2008).

    Article  CAS  Google Scholar 

  11. Correa, R. G. et al. Zebrafish IκB kinase 1 negatively regulates NF-κB activity. Curr. Biol. 15, 1291–1295 (2005).

    Article  CAS  Google Scholar 

  12. Irelan, J. T. et al. A role for IκB kinase 2 in bipolar spindle assembly. Proc. Natl Acad. Sci. USA 104, 16940–16945 (2007).

    Article  CAS  Google Scholar 

  13. Xia, Y. et al. Phosphorylation of p53 by IκB kinase 2 promotes its degradation by β-TrCP. Proc. Natl Acad. Sci. USA (2009).

  14. Chariot, A. The NF-κB-independent functions of IKK subunits in immunity and cancer. Trends Cell Biol. 19, 404–413 (2009).

    Article  CAS  Google Scholar 

  15. Perkins, N. D. Post-translational modifications regulating the activity and function of the nuclear factor κB pathway. Oncogene 25, 6717–6730 (2006).

    Article  CAS  Google Scholar 

  16. Moscat, J., Diaz-Meco, M. T. & Wooten, M. W. Signal integration and diversification through the p62 scaffold protein. Trends Biochem. Sci. 32, 95–100 (2007).

    Article  CAS  Google Scholar 

  17. Miller, K. M., Ferreira, M. G. & Cooper, J. P. Taz1, Rap1 and Rif1 act both interdependently and independently to maintain telomeres. EMBO J. 24, 3128–3135 (2005).

    Article  CAS  Google Scholar 

  18. Brevet, V. et al. The number of vertebrate repeats can be regulated at yeast telomeres by Rap1-independent mechanisms. EMBO J. 22, 1697–1706 (2003).

    Article  CAS  Google Scholar 

  19. Krauskopf, A. & Blackburn, E. H. Rap1 protein regulates telomere turnover in yeast. Proc. Natl Acad. Sci. USA 95, 12486–12491 (1998).

    Article  CAS  Google Scholar 

  20. Krauskopf, A. & Blackburn, E. H. Control of telomere growth by interactions of RAP1 with the most distal telomeric repeats. Nature 383, 354–357 (1996).

    Article  CAS  Google Scholar 

  21. Morse, R. H. RAP, RAP, open up! New wrinkles for RAP1 in yeast. Trends Genet. 16, 51–53 (2000).

    Article  CAS  Google Scholar 

  22. Yang, X., Figueiredo, L. M., Espinal, A., Okubo, E. & Li, B. RAP1 is essential for silencing telomeric variant surface glycoprotein genes in Trypanosoma brucei. Cell 137, 99–109 (2009).

    Article  CAS  Google Scholar 

  23. Li, B., Oestreich, S. & de Lange, T. Identification of human Rap1: implications for telomere evolution. Cell 101, 471–483 (2000).

    Article  CAS  Google Scholar 

  24. de Lange, T. Shelterin: the protein complex that shapes and safeguards human telomeres. Genes Dev 19, 2100–2110 (2005).

    Article  CAS  Google Scholar 

  25. Bianchi, A. & Shore, D. How telomerase reaches its end: mechanism of telomerase regulation by the telomeric complex. Mol. Cell 31, 153–165 (2008).

    Article  CAS  Google Scholar 

  26. Li, B. & de Lange, T. Rap1 affects the length and heterogeneity of human telomeres. Mol. Biol. Cell 14, 5060–5068 (2003).

    Article  CAS  Google Scholar 

  27. Bae, N. S. & Baumann, P. A RAP1/TRF2 complex inhibits nonhomologous end-joining at human telomeric DNA ends. Mol. Cell 26, 323–334 (2007).

    Article  CAS  Google Scholar 

  28. Price, C. M. wRAPing up the end to prevent telomere fusions. Mol. Cell 26, 463–464 (2007).

    Article  CAS  Google Scholar 

  29. Sfeir, A., Kabir, S., van Overbeek, M., Celli, G. B. & de Lange, T. Loss of Rap1 induces telomere recombination in the absence of NHEJ or a DNA damage signal. Science 327, 1657–1661 (2010).

    Article  CAS  Google Scholar 

  30. O'Sullivan, R. J. & Karlseder, J. Telomeres: protecting chromosomes against genome instability. Nat. Rev. Mol. Cell Biol. 11, 171–181 (2010).

    Article  CAS  Google Scholar 

  31. Gerhard, D. S. et al. The status, quality, and expansion of the NIH full-length cDNA project: the mammalian gene collection (MGC). Genome Res. 14, 2121–2127 (2004).

    Article  Google Scholar 

  32. Chanda, S. K. et al. Genome-scale functional profiling of the mammalian AP-1 signaling pathway. Proc. Natl Acad. Sci. USA 100, 12153–12158 (2003).

    Article  CAS  Google Scholar 

  33. Celli, G. B. & de Lange, T. DNA processing is not required for ATM-mediated telomere damage response after TRF2 deletion. Nat. Cell Biol. 7, 712–718 (2005).

    Article  CAS  Google Scholar 

  34. Chen, L. F. et al. NF-κB RelA phosphorylation regulates RelA acetylation. Mol. Cell Biol. 25, 7966–7975 (2005).

    Article  CAS  Google Scholar 

  35. Chen, L. F. & Greene, W. C. Shaping the nuclear action of NF-κB. Nat. Rev. Mol. Cell Biol. 5, 392–401 (2004).

    Article  CAS  Google Scholar 

  36. Natoli, G., Saccani, S., Bosisio, D. & Marazzi, I. Interactions of NF-κB with chromatin: the art of being at the right place at the right time. Nat. Immunol. 6, 439–445 (2005).

    Article  CAS  Google Scholar 

  37. Kendellen, M. F., Barrientos, K. S. & Counter, C. M. POT1 association with TRF2 regulates telomere length. Mol. Cell Biol. 29, 5611–5619 (2009).

    Article  CAS  Google Scholar 

  38. Karlseder, J. et al. Targeted deletion reveals an essential function for the telomere length regulator Trf1. Mol. Cell Biol. 23, 6533–6541 (2003).

    Article  CAS  Google Scholar 

  39. Takeuchi, O. & Akira, S. Pattern recognition receptors and inflammation. Cell 140, 805–820 (2010).

    Article  CAS  Google Scholar 

  40. Kawai, T. & Akira, S. The role of pattern-recognition receptors in innate immunity: update on Toll-like receptors. Nat. Immunol. 11, 373–384 (2010).

    Article  CAS  Google Scholar 

  41. Biswas, S. K. et al. Role for MyD88-independent, TRIF pathway in lipid A/TLR4-induced endotoxin tolerance. J. Immunol. 179, 4083–4092 (2007).

    Article  CAS  Google Scholar 

  42. Weaver, V. M. et al. β4 integrin-dependent formation of polarized three-dimensional architecture confers resistance to apoptosis in normal and malignant mammary epithelium. Cancer Cell 2, 205–216 (2002).

    Article  CAS  Google Scholar 

  43. Sovak, M. A. et al. Aberrant nuclear factor-κB/Rel expression and the pathogenesis of breast cancer. J. Clin. Invest. 100, 2952–2960 (1997).

    Article  CAS  Google Scholar 

  44. Weigelt, B., Peterse, J. L. & van' t Veer, L. J. Breast cancer metastasis: markers and models. Nat. Rev. Cancer 5, 591–602 (2005).

    Article  CAS  Google Scholar 

  45. Basseres, D. S. & Baldwin, A. S. Nuclear factor-κB and inhibitor of κB kinase pathways in oncogenic initiation and progression. Oncogene 25, 6817–6830 (2006).

    Article  CAS  Google Scholar 

  46. Dey, A., Wong, E. T., Cheok, C. F., Tergaonkar, V. & Lane, D. P. R-Roscovitine simultaneously targets both the p53 and NF-κB pathways and causes potentiation of apoptosis: implications in cancer therapy. Cell Death Differ. 15, 263–273 (2008).

    Article  CAS  Google Scholar 

  47. Dey, A., Wong, E. T., Bist, P., Tergaonkar, V. & Lane, D. P. Nutlin-3 inhibits the NFκB pathway in a p53-dependent manner: implications in lung cancer therapy. Cell Cycle 6, 2178–2185 (2007).

    Article  CAS  Google Scholar 

  48. Papa, S. et al. The NF-κB-mediated control of the JNK cascade in the antagonism of programmed cell death in health and disease. Cell Death Differ. 13, 712–729 (2006).

    Article  CAS  Google Scholar 

  49. Chua, H. L. et al. NF-κB represses E-cadherin expression and enhances epithelial to mesenchymal transition of mammary epithelial cells: potential involvement of ZEB-1 and ZEB-2. Oncogene 26, 711–724 (2007).

    Article  CAS  Google Scholar 

  50. Campbell, M. J. et al. Breast cancer growth prevention by statins. Cancer Res. 66, 8707–8714 (2006).

    Article  CAS  Google Scholar 

  51. Biswas, D. K. et al. NF-κB activation in human breast cancer specimens and its role in cell proliferation and apoptosis. Proc. Natl Acad. Sci. USA 101, 10137–10142 (2004).

    Article  CAS  Google Scholar 

  52. Micheau, O. & Tschopp, J. Induction of TNF receptor I-mediated apoptosis via two sequential signaling complexes. Cell 114, 181–190 (2003).

    Article  CAS  Google Scholar 

  53. Helbig, G. et al. NF-κB promotes breast cancer cell migration and metastasis by inducing the expression of the chemokine receptor CXCR4. J. Biol. Chem. 278, 21631–21638 (2003).

    Article  CAS  Google Scholar 

  54. Rao Ch, V., Li, X., Manna, S. K., Lei, Z. M. & Aggarwal, B. B. Human chorionic gonadotropin decreases proliferation and invasion of breast cancer MCF-7 cells by inhibiting NF-κB and AP-1 activation. J. Biol. Chem. 279, 25503–25510 (2004).

    Article  CAS  Google Scholar 

  55. Hagemann, T. et al. Macrophages induce invasiveness of epithelial cancer cells via NF-κB and JNK. J. Immunol. 175, 1197–1205 (2005).

    Article  CAS  Google Scholar 

  56. Chew, J. et al. WIP1 phosphatase is a negative regulator of NF-κB signalling. Nat. Cell Biol. 11, 659–666 (2009).

    Article  CAS  Google Scholar 

  57. Park, J. I. et al. Telomerase modulates Wnt signalling by association with target gene chromatin. Nature 460, 66–72 (2009).

    Article  CAS  Google Scholar 

  58. Huang, S. M. et al. Tankyrase inhibition stabilizes axin and antagonizes Wnt signalling. Nature 461, 614–620 (2009).

    Article  CAS  Google Scholar 

  59. Tergaonkar, V., Bottero, V., Ikawa, M., Li, Q. & Verma, I. M. IκB kinase-independent IκBα degradation pathway: functional NF-κB activity and implications for cancer therapy. Mol. Cell Biol. 23, 8070–8083 (2003).

    Article  CAS  Google Scholar 

  60. Deng, Z., Atanasiu, C., Burg, J. S., Broccoli, D. & Lieberman, P. M. Telomere repeat binding factors TRF1, TRF2, and hRAP1 modulate replication of Epstein-Barr virus OriP. J. Virol. 77, 11992–12001 (2003).

    Article  CAS  Google Scholar 

  61. Cho, C. Y. et al. Identification of the tyrosine phosphatase PTP-MEG2 as an antagonist of hepatic insulin signaling. Cell Metab. 3, 367–378 (2006).

    Article  CAS  Google Scholar 

  62. Luesch, H. et al. A functional genomics approach to the mode of action of apratoxin A. Nat. Chem. Biol. 2, 158–167 (2006).

    Article  CAS  Google Scholar 

  63. Yu, F. et al. Systematic identification of cellular signals reactivating Kaposi sarcoma-associated herpesvirus. PLoS Pathog. 3, e44 (2007).

    Article  Google Scholar 

Download references

Acknowledgements

We wish to thank the Agency for Science Technology and Research, Singapore (A*Star) for funding and support to the V.T. laboratory. I.M.V. is an American Cancer Society Professor of Molecular Biology, and holds the Joan and Irwin Jacobs Chair in Exemplary Life Sciences. This work was supported in part by grants from the Leducq Foundation, Meriaux Foundation, Ellison Medical Foundation, Ipsen/Biomeasure, Sanofi Aventis, and the H.N. and Frances C. Berger Foundation. We thank T. Murphy and N. Tonnu for help with the intial investigations involved in this work. We are grateful to B. Li and T. De Lange for Rap1 deletion constructs and P. Lieberman for the Rap1 shRNA vector. We thank J. Karlseder for helpful discussions.

Author information

Author notes

  1. Sourav Ghosh, Hendrik Luesch, Arkasubhra Ghosh and Ee Tsin Wong: These authors contributed equally to this work.

Authors and Affiliations

  1. Laboratory of NFκB Signaling, Proteos, 138673, Singapore, Singapore

    Hsiangling Teo, Arkasubhra Ghosh, Ee Tsin Wong, Najib Malik, Marc Wong & Vinay Tergaonkar

  2. Department of Basic Medical Sciences, University of Arizona College of Medicine, Phoenix, 85440, AZ, USA

    Sourav Ghosh & Lisa J. Speiser

  3. Department of Medicinal Chemistry, University of Florida, Gainesville, 32610, FL, USA

    Hendrik Luesch

  4. Genomics Institute of the Novartis Research Foundation, San Diego, 92121, CA, USA

    Hendrik Luesch, Anthony Orth, Paul de Jesus, Enrique Saez, Peter Schultz & Sumit K. Chanda

  5. Department of Pathology, St. Joseph's Hospital and Medical Center, Phoenix, 85004, AZ, USA

    Anthony S. Perry & Jeffrey D. Oliver

  6. CNS Tumour Research Laboratory, Translational Genomics Research Institute, Phoenix, 85004, AZ, USA

    Nhan L. Tran

  7. Infectious and Inflammatory Disease Center, Sanford-Burnham Medical Research Institute, La Jolla, 92037, CA, USA

    Sumit K. Chanda

  8. Laboratory of genetics, Salk Institute for Biological Studies, La Jolla, 92037, CA, USA

    Inder M. Verma

Authors
  1. Hsiangling Teo
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Sourav Ghosh
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Hendrik Luesch
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Arkasubhra Ghosh
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Ee Tsin Wong
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Najib Malik
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Anthony Orth
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Paul de Jesus
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Anthony S. Perry
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. Jeffrey D. Oliver
    View author publications

    You can also search for this author in PubMed Google Scholar

  11. Nhan L. Tran
    View author publications

    You can also search for this author in PubMed Google Scholar

  12. Lisa J. Speiser
    View author publications

    You can also search for this author in PubMed Google Scholar

  13. Marc Wong
    View author publications

    You can also search for this author in PubMed Google Scholar

  14. Enrique Saez
    View author publications

    You can also search for this author in PubMed Google Scholar

  15. Peter Schultz
    View author publications

    You can also search for this author in PubMed Google Scholar

  16. Sumit K. Chanda
    View author publications

    You can also search for this author in PubMed Google Scholar

  17. Inder M. Verma
    View author publications

    You can also search for this author in PubMed Google Scholar

  18. Vinay Tergaonkar
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

H.T., A.G., E.T.W., N.M., M.W. and V.T. carried out biochemical experiments with Rap1, generated Rap1-mutant mice and helped in data analysis. S.G., A.S.P., J.D.O., N.L.T. and L.J.S. carried out the analysis of Rap1 in human cancers and also helped in data analysis. H.L., P.D.J., A.O., E.S., P.S., S.K.C. and V.T. carried out the genome-wide screen and helped in data analysis and initial characterization of Rap1 as an NF-κB regulator. S.K.C., I.M.V. and V.T. did the project planning and data analysis.

Corresponding author

Correspondence to Vinay Tergaonkar.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Information

Supplementary Information (PDF 1112 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Teo, H., Ghosh, S., Luesch, H. et al. Telomere-independent Rap1 is an IKK adaptor and regulates NF-κB-dependent gene expression. Nat Cell Biol 12, 758–767 (2010). https://doi.org/10.1038/ncb2080

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/ncb2080

This article is cited by

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing