Identification of a co-activator that links growth factor signalling to c-Jun/AP-1 activation


The AP-1 transcription factor c-Jun is essential for cellular proliferation in many cell types, but the molecular link between growth factors and c-Jun activation has been enigmatic. In this study we identify a previously uncharacterized RING-domain-containing protein, RACO-1 (RING domain AP-1 co-activator-1), as a c-Jun co-activator that is regulated by growth factor signalling. RACO-1 interacted with c-Jun independently of amino-terminal phosphorylation, and was both necessary and sufficient for c-Jun/AP-1 activation. Growth factor-mediated stimulation of AP-1 was attributable to MEK/ERK-dependent stabilization of RACO-1 protein. Stimulation of the MEK/ERK pathway strongly promoted Lys 63-linked ubiquitylation of RACO-1, which antagonized Lys 48-linked degradative auto-ubiquitylation of the same Lys residues. RACO-1 depletion reduced cellular proliferation and decreased expression of several growth-associated AP-1 target genes, such as cdc2, cyclinD1 and hb-egf. Moreover, transgenic overexpression of RACO-1 augmented intestinal tumour formation triggered by aberrant Wnt signalling and cooperated with oncogenic Ras in colonic hyperproliferation. Thus RACO-1 is a co-activator that links c-Jun to growth factor signalling and is essential for AP-1 function in proliferation.

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Figure 1: RACO-1 binds to c-Jun independently of N-terminal phosphorylation.
Figure 2: RACO-1 is a MEKK1-dependent c-Jun co-activator regulated by MAP kinase signalling pathways.
Figure 3: Growth factor signalling through the MEK pathway stabilizes RACO-1 protein.
Figure 4: MEK signalling promotes the formation of Lys 63-linked ubiquitin chains that antagonize degradative auto-ubiquitylation.
Figure 5: The residues targeted for MEK1-induced ubiquitylation chain elongation are also critical for degradative poly-ubiquitylation.
Figure 6: RACO-1 is an important regulator of cell-cycle progression and proliferation in NIH3T3 fibroblasts.
Figure 7: Transgenic expression of RACO-1 in vivo enhances APCmin+-induced intestinal tumorigenesis and cooperates with oncogenic Ras in colonic hyperproliferation.


  1. 1

    Shaulian, E. & Karin, M. AP-1 in cell proliferation and survival. Oncogene 20, 2390–2400 (2001).

    CAS  Article  Google Scholar 

  2. 2

    Eferl, R. & Wagner, E. F. AP-1: a double-edged sword in tumorigenesis. Nature Rev. Cancer 3, 859–868 (2003).

    CAS  Article  Google Scholar 

  3. 3

    Mechta-Grigoriou, F., Gerald, D. & Yaniv, M. The mammalian Jun proteins: redundancy and specificity. Oncogene 20, 2378–2389 (2001).

    CAS  Article  Google Scholar 

  4. 4

    Davis, R. J. Signal transduction by the JNK group of MAP kinases. Cell 103, 239–252 (2000).

    CAS  Article  Google Scholar 

  5. 5

    Angel, P., Hattori, K., Smeal, T. & Karin, M. The jun proto-oncogene is positively autoregulated by its product, Jun/AP1. Cell 55, 875–885 (1988).

    CAS  Article  Google Scholar 

  6. 6

    Chen, Z. et al. MAP kinases. Chem. Rev. 101, 2449–2476 (2001).

    CAS  Article  Google Scholar 

  7. 7

    Pearson, G. et al. Mitogen-activated protein (MAP) kinase pathways: regulation and physiological functions. Endocr. Rev. 22, 153–183 (2001).

    CAS  PubMed  Google Scholar 

  8. 8

    Johnson, R. S., van Lingen, B., Papaioannou, V. E. & Spiegelmann, B. M. A null mutation at the c-jun locus causes embryonic lethality and retarded cell growth in culture. Genes Dev. 7, 1309–1317 (1993).

    CAS  Article  Google Scholar 

  9. 9

    Schreiber, M. et al. Control of cell cycle progression by c-Jun is p53 dependent. Genes Dev. 13, 607–619 (1999).

    CAS  Article  Google Scholar 

  10. 10

    Behrens, A., Sibilia, M. & Wagner, E. F. Amino-terminal phosphorylation of c-Jun regulates stress-induced apoptosis and cellular proliferation. Nature Genet. 21, 326–329 (1999).

    CAS  Article  Google Scholar 

  11. 11

    Eferl, R. et al. Liver tumor development. c-Jun antagonizes the proapoptotic activity of p53. Cell 112, 181–192 (2003).

    CAS  Article  Google Scholar 

  12. 12

    Hilberg, F., Aguzzi, A., Howells, N. & Wagner, E. F. c-jun is essential for normal mouse development and hepatogenesis. Nature 365, 179–181 (1993).

    CAS  Article  Google Scholar 

  13. 13

    Behrens, A. et al. Impaired intervertebral disc formation in the absence of Jun. Development 130, 103–109 (2003).

    CAS  Article  Google Scholar 

  14. 14

    Eferl, R. et al. Functions of c-Jun in liver and heart development. J. Cell Biol. 145, 1049–1061 (1999).

    CAS  Article  Google Scholar 

  15. 15

    Behrens, A., Jochum, W., Sibilia, M. & Wagner, E. F. Oncogenic transformation by ras and fos is mediated by c-Jun N-terminal phosphorylation. Oncogene 19, 2657–2663 (2000).

    CAS  Article  Google Scholar 

  16. 16

    Lee, W., Mitchell, P. & Tjian, R. Purified transcription factor AP-1 interacts with TPA-inducible enhancer elements. Cell 49, 741–752 (1987).

    CAS  Article  Google Scholar 

  17. 17

    Angel, P. et al. Phorbol ester-inducible genes contain a common cis element recognized by a TPA-modulated trans-acting factor. Cell 49, 729–739 (1987).

    CAS  Article  Google Scholar 

  18. 18

    Morton, S., Davis, R. J., McLaren, A. & Cohen, P. A reinvestigation of the multisite phosphorylation of the transcription factor c-Jun. EMBO J. 22, 3876–3886 (2003).

    CAS  Article  Google Scholar 

  19. 19

    Nateri, A. S., Riera-Sans, L., Da Costa, C. & Behrens, A. The ubiquitin ligase SCFFbw7 antagonizes apoptotic JNK signaling. Science 303, 1374–1378 (2004).

    CAS  Article  Google Scholar 

  20. 20

    Ardley, H. C. & Robinson, P. A. E3 ubiquitin ligases. Essays Biochem. 41, 15–30 (2005).

    CAS  Article  Google Scholar 

  21. 21

    Fang, D. et al. Dysregulation of T lymphocyte function in itchy mice: a role for Itch in TH2 differentiation. Nature Immunol. 3, 281–287 (2002).

    CAS  Article  Google Scholar 

  22. 22

    Gao, M. et al. Jun turnover is controlled through JNK-dependent phosphorylation of the E3 ligase Itch. Science 306, 271–275 (2004).

    CAS  Article  Google Scholar 

  23. 23

    Wei, W., Jin, J., Schlisio, S., Harper, J. W. & Kaelin, W. G., Jr. The v-Jun point mutation allows c-Jun to escape GSK3-dependent recognition and destruction by the Fbw7 ubiquitin ligase. Cancer Cell 8, 25–33 (2005).

    CAS  Article  Google Scholar 

  24. 24

    Wertz, I. E et al. Human De-etiolated-1 regulates c-Jun by assembling a CUL4A ubiquitin ligase. Science 303, 1371–1374 (2004).

    CAS  Article  Google Scholar 

  25. 25

    De Cesare, D. et al. Heterodimerization of c-Jun with ATF-2 and c-Fos is required for positive and negative regulation of the human urokinase enhancer. Oncogene 11, 365–376 (1995).

    CAS  PubMed  Google Scholar 

  26. 26

    Nateri, A. S., Spencer-Dene, B. & Behrens, A. Interaction of phosphorylated c-Jun with TCF4 regulates intestinal cancer development. Nature 437, 281–285 (2005).

    CAS  Article  Google Scholar 

  27. 27

    Wada, T. et al. MKK7 couples stress signalling to G2/M cell-cycle progression and cellular senescence. Nature Cell Biol. 6, 215–226 (2004).

    CAS  Article  Google Scholar 

  28. 28

    Yujiri, T., Sather, S., Fanger, G. R. & Johnson, G. L. Role of MEKK1 in cell survival and activation of JNK and ERK pathways defined by targeted gene disruption. Science 282, 1911–1914 (1998).

    CAS  Article  Google Scholar 

  29. 29

    Mansour, S. J., Candia, J. M., Gloor, K. K. & Ahn, N. G. Constitutively active mitogen-activated protein kinase kinase 1 (MAPKK1) and MAPKK2 mediate similar transcriptional and morphological responses. Cell Growth Differ. 7, 243–250 (1996).

    CAS  PubMed  PubMed Central  Google Scholar 

  30. 30

    Wisdom, R., Johnson, R. S. & Moore, C. c-Jun regulates cell cycle progression and apoptosis by distinct mechanisms. Embo J. 18, 188–197 (1999).

    CAS  Article  Google Scholar 

  31. 31

    Nyabi, O. et al. Efficient mouse transgenesis using Gateway-compatible ROSA26 locus targeting vectors and F1 hybrid ES cells. Nucleic Acids Res. 37, e55 (2009).

    Article  Google Scholar 

  32. 32

    el Marjou, F. et al. Tissue-specific and inducible Cre-mediated recombination in the gut epithelium. Genesis 39, 186–193 (2004).

    CAS  Article  Google Scholar 

  33. 33

    D'Abaco, G. M., Whitehead, R. H. & Burgess, A. W. Synergy between Apc min and an activated ras mutation is sufficient to induce colon carcinomas. Mol. Cell Biol. 16, 884–891 (1996).

    CAS  Article  Google Scholar 

  34. 34

    Janssen, K. P. et al. APC and oncogenic KRAS are synergistic in enhancing Wnt signaling in intestinal tumor formation and progression. Gastroenterology 131, 1096–1109 (2006).

    CAS  Article  Google Scholar 

  35. 35

    Tuveson, D. A. et al. Endogenous oncogenic K-ras(G12D) stimulates proliferation and widespread neoplastic and developmental defects. Cancer Cell 5, 375–387 (2004).

    CAS  Article  Google Scholar 

  36. 36

    Bohmann, D. et al. Human proto-oncogene c-jun encodes a DNA binding protein with structural and functional properties of transcription factor AP-1. Science 238, 1386–1392 (1987).

    CAS  Article  Google Scholar 

  37. 37

    Treisman, R. Journey to the surface of the cell: Fos regulation and the SRE. EMBO J. 14, 4905–4913 (1995).

    CAS  Article  Google Scholar 

  38. 38

    Hu, E. et al. Targeted disruption of the c-fos gene demonstrates c-fos-dependent and -independent pathways for gene expression stimulated by growth factors or oncogenes. EMBO J. 13, 3094–3103 (1994).

    CAS  Article  Google Scholar 

  39. 39

    Shaulian, E. et al. The mammalian UV response: c-Jun induction is required for exit from p53-imposed growth arrest. Cell 103, 897–907 (2000).

    CAS  Article  Google Scholar 

  40. 40

    Johnson, R., Spiegelman, B., Hanahan, D. & Wisdom, R. Cellular transformation and malignancy induced by ras require c-jun. Mol. Cell Biol. 16, 4504–4511 (1996).

    CAS  Article  Google Scholar 

  41. 41

    Minden, A., Lin, A., Claret, F. X., Abo, A. & Karin, M. Selective activation of the JNK signaling cascade and c-Jun transcriptional activity by the small GTPases Rac and Cdc42Hs. Cell 81, 1147–1157 (1995).

    CAS  Article  Google Scholar 

  42. 42

    de Ruiter, N. D., Wolthuis, R. M., van Dam, H., Burgering, B. M. & Bos, J. L. Ras-dependent regulation of c-Jun phosphorylation is mediated by the Ral guanine nucleotide exchange factor-Ral pathway. Mol. Cell Biol. 20, 8480–8488 (2000).

    CAS  Article  Google Scholar 

  43. 43

    Kennedy, N. J. et al. Suppression of Ras-stimulated transformation by the JNK signal transduction pathway. Genes Dev. 17, 629–637 (2003).

    CAS  Article  Google Scholar 

  44. 44

    Alberts, A. S., Geneste, O. & Treisman, R. Activation of SRF-regulated chromosomal templates by Rho-family GTPases requires a signal that also induces H4 hyperacetylation. Cell 92, 475–487 (1998).

    CAS  Article  Google Scholar 

  45. 45

    Campanero, M. R. & Flemington, E. K. Regulation of E2F through ubiquitin-proteasome-dependent degradation: stabilization by the pRB tumor suppressor protein. Proc. Natl Acad. Sci. USA 94, 2221–2226 (1997).

    CAS  Article  Google Scholar 

  46. 46

    George, S. H. et al. Developmental and adult phenotyping directly from mutant embryonic stem cells. Proc. Natl Acad. Sci. USA 104, 4455–4460 (2007).

    CAS  Article  Google Scholar 

  47. 47

    Sancho, R. et al. JNK signalling modulates intestinal homeostasis and tumourigenesis in mice. EMBO J. 28, 1843–1854 (2009).

    CAS  Article  Google Scholar 

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We are grateful to N. Ahn, M. Cobb and A. Hock for providing reagents. We thank B. Spencer-Dean, E. Nye and R. Mitter for technical support; the Molecular Biology Core Facility, Paterson Institute for Cancer Research, Manchester, UK for microarray analysis; and H. Walden for critical reading of the manuscript. The London Research Institute is funded by Cancer Research UK.

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C.D and A.C performed all experiments; C.D and A.B designed and analysed all experiments and wrote the manuscript; K.H and J.H generated R26RACO−1 transgenic mice; F.C generated RACO-1 antibodies.

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Correspondence to Axel Behrens.

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The authors declare no competing financial interests.

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Davies, C., Chakraborty, A., Cipriani, F. et al. Identification of a co-activator that links growth factor signalling to c-Jun/AP-1 activation. Nat Cell Biol 12, 963–972 (2010).

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