Abstract
Signalling activity of the Notch receptor, which plays a fundamental role in metazoan cell fate determination, is controlled at multiple levels. We uncovered a Notch signal-controlling mechanism that depends on the ability of the non-visual β-arrestin, Kurtz (Krz), to influence the degradation and, consequently, the function of the Notch receptor. We identified Krz as a binding partner of a known Notch-pathway modulator, Deltex (Dx), and demonstrated the existence of a trimeric Notch–Dx–Krz protein complex. This complex mediates the degradation of the Notch receptor through a ubiquitination-dependent pathway. Our results establish a novel mode of regulation of Notch signalling and define a new function for non-visual β-arrestins.
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Artavanis-Tsakonas, S., Rand, M. D. & Lake, R. J. Notch signaling: cell fate control and signal integration in development. Science 284, 770–776 (1999).
Harper, J. A., Yuan, J. S., Tan, J. B., Visan, I. & Guidos, C. J. Notch signaling in development and disease. Clin. Genet. 64, 461–472 (2003).
Ramos, R. G., Grimwade, B. G., Wharton, K. A., Scottgale, T. N. & Artavanis-Tsakonas, S. Physical and functional definition of the Drosophila Notch locus by P element transformation. Genetics 123, 337–348 (1989).
Roman, G., He, J. & Davis, R. L. kurtz, a novel nonvisual arrestin, is an essential neural gene in Drosophila. Genetics 155, 1281–1295 (2000).
Busseau, I., Diederich, R. J., Xu, T. & Artavanis-Tsakonas, S. A member of the Notch group of interacting loci, deltex encodes a cytoplasmic basic protein. Genetics 136, 585–596 (1994).
Diederich, R. J., Matsuno, K., Hing, H. & Artavanis-Tsakonas, S. Cytosolic interaction between deltex and Notch ankyrin repeats implicates deltex in the Notch signaling pathway. Development 120, 473–481 (1994).
Aravind, L. The WWE domain: a common interaction module in protein ubiquitination and ADP ribosylation. Trends Biochem. Sci. 26, 273–275 (2001).
Matsuno, K., Diederich, R. J., Go, M. J., Blaumueller, C. M. & Artavanis-Tsakonas, S. Deltex acts as a positive regulator of Notch signaling through interactions with the Notch ankyrin repeats. Development 121, 2633–2644 (1995).
Takeyama, K. et al. The BAL-binding protein BBAP and related Deltex family members exhibit ubiquitin-protein isopeptide ligase activity. J. Biol. Chem. 278, 21930–21937 (2003).
Lefkowitz, R. J. & Shenoy, S. K. Transduction of receptor signals by β-arrestins. Science 308, 512–517 (2005).
Luttrell, L. M. & Lefkowitz, R. J. The role of β-arrestins in the termination and transduction of G-protein-coupled receptor signals. J. Cell Sci. 115, 455–465 (2002).
Chen, W. et al. β-Arrestin 2 mediates endocytosis of type III TGF-β receptor and down-regulation of its signaling. Science 301, 1394–1397 (2003).
Chen, W. et al. Dishevelled 2 recruits β-arrestin 2 to mediate Wnt5A-stimulated endocytosis of Frizzled 4. Science 301, 1391–1394 (2003).
Chen, W. et al. Activity-dependent internalization of smoothened mediated by β-arrestin 2 and GRK2. Science 306, 2257–2260 (2004).
Lin, F. T., Daaka, Y. & Lefkowitz, R. J. β-Arrestins regulate mitogenic signaling and clathrin-mediated endocytosis of the insulin-like growth factor I receptor. J. Biol. Chem. 273, 31640–31643 (1998).
Wilbanks, A. M. et al. β-arrestin 2 regulates zebrafish development through the hedgehog signaling pathway. Science 306, 2264–2267 (2004).
Wu, J. H. et al. The adaptor protein β-arrestin2 enhances endocytosis of the low density lipoprotein receptor. J. Biol. Chem. 278, 44238–44245 (2003).
Gao, H. et al. Identification of β-arrestin2 as a G protein-coupled receptor-stimulated regulator of NF-κB pathways. Mol. Cell 14, 303–317 (2004).
Witherow, D. S., Garrison, T. R., Miller, W. E. & Lefkowitz, R. J. β-Arrestin inhibits NF-κB activity by means of its interaction with the NF-κB inhibitor IκBα. Proc. Natl Acad. Sci. USA 101, 8603–8607 (2004).
Fromont-Racine, M., Rain, J. C. & Legrain, P. Toward a functional analysis of the yeast genome through exhaustive two-hybrid screens. Nature Genet. 16, 277–282 (1997).
Veraksa, A., Bauer, A. & Artavanis-Tsakonas, S. Analyzing protein complexes in Drosophila with tandem affinity purification-mass spectrometry. Dev. Dyn. 232, 827–834 (2005).
Xu, T. & Artavanis-Tsakonas, S. Deltex, a locus interacting with the neurogenic genes, Notch, Delta and mastermind in Drosophila melanogaster. Genetics 126, 665–677 (1990).
Fehon, R. G. et al. Molecular interactions between the protein products of the neurogenic loci Notch and Delta, two EGF-homologous genes in Drosophila. Cell 61, 523–534 (1990).
Xu, T. & Rubin, G. M. Analysis of genetic mosaics in developing and adult Drosophila tissues. Development 117, 1223–1237 (1993).
de Celis, J. F., Garcia-Bellido, A. & Bray, S. J. Activation and function of Notch at the dorsal-ventral boundary of the wing imaginal disc. Development 122, 359–369 (1996).
Neumann, C. J. & Cohen, S. M. A hierarchy of cross-regulation involving Notch, wingless, vestigial and cut organizes the dorsal/ventral axis of the Drosophila wing. Development 122, 3477–3485 (1996).
Tsuda, L., Nagaraj, R., Zipursky, S. L. & Banerjee, U. An EGFR/Ebi/Sno pathway promotes delta expression by inactivating Su(H)/SMRTER repression during inductive notch signaling. Cell 110, 625–637 (2002).
Hori, K. et al. Drosophila Deltex mediates Suppressor of Hairless-independent and late-endosomal activation of Notch signaling. Development 131, 5527–5537 (2004).
Shenoy, S. K., McDonald, P. H., Kohout, T. A. & Lefkowitz, R. J. Regulation of receptor fate by ubiquitination of activated β2-adrenergic receptor and β-arrestin. Science 294, 1307–1313 (2001).
Girnita, L. et al. β-Arrestin is crucial for ubiquitination and down-regulation of the insulin-like growth factor-1 receptor by acting as adaptor for the MDM2 E3 ligase. J. Biol. Chem. 280, 24412–24419 (2005).
Matsuno, K. et al. Human deltex is a conserved regulator of Notch signalling. Nature Genet. 19, 74–78 (1998).
Yamamoto, N. et al. Role of Deltex-1 as a transcriptional regulator downstream of the Notch receptor. J. Biol. Chem. 276, 45031–45040 (2001).
Ordentlich, P. et al. Notch inhibition of E47 supports the existence of a novel signaling pathway. Mol. Cell. Biol. 18, 2230–2239 (1998).
Ramain, P. et al. Novel Notch alleles reveal a Deltex-dependent pathway repressing neural fate. Curr. Biol. 11, 1729–1738 (2001).
Izon, D. J. et al. Deltex1 redirects lymphoid progenitors to the B cell lineage by antagonizing Notch1. Immunity 16, 231–243 (2002).
Kiaris, H. et al. Modulation of notch signaling elicits signature tumors and inhibits hras1-induced oncogenesis in the mouse mammary epithelium. Am. J. Pathol. 165, 695–705 (2004).
Sestan, N., Artavanis-Tsakonas, S. & Rakic, P. Contact-dependent inhibition of cortical neurite growth mediated by notch signaling. Science 286, 741–746 (1999).
Matsuno, K. et al. Involvement of a proline-rich motif and RING-H2 finger of Deltex in the regulation of Notch signaling. Development 129, 1049–1059 (2002).
Shenoy, S. K. & Lefkowitz, R. J. Trafficking patterns of β-arrestin and G protein-coupled receptors determined by the kinetics of β-arrestin deubiquitination. J. Biol. Chem. 278, 14498–14506 (2003).
Gupta-Rossi, N. et al. Monoubiquitination and endocytosis direct γ-secretase cleavage of activated Notch receptor. J. Cell Biol. 166, 73–83 (2004).
Oberg, C. et al. The Notch intracellular domain is ubiquitinated and negatively regulated by the mammalian Sel-10 homolog. J. Biol. Chem. 276, 35847–35853 (2001).
Wu, G. et al. SEL-10 is an inhibitor of notch signaling that targets notch for ubiquitin-mediated protein degradation. Mol. Cell. Biol. 21, 7403–7415 (2001).
Cornell, M. et al. The Drosophila melanogaster Suppressor of deltex gene, a regulator of the Notch receptor signaling pathway, is an E3 class ubiquitin ligase. Genetics 152, 567–576 (1999).
Wilkin, M. B. et al. Regulation of notch endosomal sorting and signaling by Drosophila Nedd4 family proteins. Curr. Biol. 14, 2237–2244 (2004).
Sakata, T. et al. Drosophila Nedd4 regulates endocytosis of notch and suppresses its ligand-independent activation. Curr. Biol. 14, 2228–2236 (2004).
Weng, A. P. et al. Activating mutations of NOTCH1 in human T cell acute lymphoblastic leukemia. Science 306, 269–271 (2004).
Bouwmeester, T. et al. A physical and functional map of the human TNF-α/NF-κB signal transduction pathway. Nature Cell Biol. 6, 97–105 (2004).
Rebay, I., Fehon, R. G. & Artavanis-Tsakonas, S. Specific truncations of Drosophila Notch define dominant activated and dominant negative forms of the receptor. Cell 74, 319–329 (1993).
Nellesen, D. T., Lai, E. C. & Posakony, J. W. Discrete enhancer elements mediate selective responsiveness of enhancer of split complex genes to common transcriptional activators. Dev. Biol. 213, 33–53 (1999).
Naidoo, N., Song, W., Hunter-Ensor, M. & Sehgal, A. A role for the proteasome in the light response of the timeless clock protein. Science 285, 1737–1741 (1999).
Acknowledgements
The authors wish to thank their colleagues D. Finley, M. Baron, R. Lake, M. Kankel and L. Grimm for helpful discussions about the manuscript. We are also grateful to M. Gonzalez-Gaitan and H. Chang for their comments and reagents. krz alleles were kindly provided by R. Davis and the dx152 allele was a gift from K. Matsuno. The anti-EGFR antibody was a gift from P. Rorth. R. Fehon provided a ubiquitination assay protocol and an HS–HA–Ub construct. pMT–Flag–UbWT and pMT–Flag–UbMONO were a gift from S. Bray. The GFP–Sec61 construct was a gift from G. Voeltz. Several fly lines were provided by the Bloomington Drosophila Stock Center, and some of the antibodies used in this work were obtained from the Developmental Studies Hybridoma Bank, Iowa. This work was supported by grants NS26084, GM62931 and CA098402 to S.A.-T. A.V. was supported by a postdoctoral fellowship from the Massachusetts General Hospital Fund for Medical Discovery.
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Mukherjee, A., Veraksa, A., Bauer, A. et al. Regulation of Notch signalling by non-visual β-arrestin. Nat Cell Biol 7, 1191–1201 (2005). https://doi.org/10.1038/ncb1327
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DOI: https://doi.org/10.1038/ncb1327
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