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Differential subcellular localization regulates c-Cbl E3 ligase activity upon Notch3 protein in T-cell leukemia

Oncogene volume 29, pages 1463–1474 (2010)Cite this article

Abstract

Notch3 and pTα signaling events are essential for T-cell leukemogenesis and characterize murine and human T-cell acute lymphoblastic leukemia. Genetic ablation of pTα expression in Notch3 transgenic mice abrogates tumor development, indicating that pTα signaling is crucial to the Notch3-mediated leukemogenesis. Here we report a novel direct interaction between Notch3 and pTα. This interaction leads to the recruitment and persistence of the E3 ligase protein c-Cbl to the lipid rafts in Notch3-IC transgenic thymocytes. Conversely, deletion of pTα in Notch3 transgenic mice leads to cytoplasmic retention of c-Cbl that targets Notch3 protein to the proteasomal-degradative pathway. It appears that protein kinase C θ (PKCθ), by regulating tyrosine and serine phosphorylation of Cbl, is able to control its function. We report here that the increased Notch3-IC degradation correlates with higher levels of c-Cbl tyrosine phosphorylation in Notch3-IC/pTα−/− double-mutant thymocytes, which also display a decreased PKCθ activity. Our data indicate that pTα/pre-T-cell receptor is able to regulate the different subcellular localization of c-Cbl and, by regulating PKCθ activity, is also able to influence its ubiquitin ligase activity upon Notch3 protein.

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References

  • Aifantis I, Azogui O, Feinberg J, Saint-Ruf C, Buer J, von Boehmer H . (1998). On the role of the pre-T cell receptor in alphabeta versus gammadelta T lineage commitment. Immunity 9: 649–655.

    Article  CAS  PubMed  Google Scholar 

  • Aifantis I, Gounari F, Scorrano L, Borowski C, von Boehmer H . (2001). Constitutive pre-TCR signaling promotes differentiation through Ca2+ mobilization and activation of NF-kappaB and NFAT. Nat Immunol 2: 403–409.

    Article  CAS  PubMed  Google Scholar 

  • Artavanis-Tsakonas S, Rand MD, Lake RJ . (1999). Notch signaling: cell fate control and signal integration in development. Science 284: 770–776.

    Article  CAS  PubMed  Google Scholar 

  • Bellavia D, Campese AF, Alesse E, Vacca A, Felli MP, Balestri A et al. (2000). Constitutive activation of NF-kappaB and T-cell leukemia/lymphoma in Notch3 transgenic mice. EMBO J 19: 3337–3348.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Bellavia D, Campese AF, Checquolo S, Balestri A, Biondi A, Cazzaniga G et al. (2002). Combined expression of pTalpha and Notch3 in T cell leukemia identifies the requirement of preTCR for leukemogenesis. Proc Natl Acad Sci USA 99: 3788–3793.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Bellavia D, Mecarozzi M, Campese AF, Grazioli P, Talora C, Frati L et al. (2007). Notch3 and the Notch3-upregulated RNA-binding protein HuD regulate Ikaros alternative splicing. EMBO J 26: 1670–1680.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Benson RA, Adamson K, Corsin-Jimenez M, Marley JV, Wahl KA, Lamb JR et al. (2005). Notch1 co-localizes with CD4 on activated T cells and Notch signaling is required for IL-10 production. Eur J Immunol 35: 859–869.

    Article  CAS  PubMed  Google Scholar 

  • Blaumueller CM, Qi H, Zagouras P, Artavanis-Tsakonas S . (1997). Intracellular cleavage of Notch leads to a heterodimeric receptor on the plasma membrane. Cell 90: 281–291.

    Article  CAS  PubMed  Google Scholar 

  • Brizzi MF, Rosso A, Dentelli P, Ferrero D, Lanfrancone L, Pegoraro L . (1998). c-Cbl tyrosine phosphorylation and subcellular localization in human primary leukemic cells. Exp Hematol 26: 1229–1239.

    CAS  PubMed  Google Scholar 

  • Campese AF, Bellavia D, Gulino A, Screpanti I . (2003). Notch signalling at the crossroads of T cell development and leukemogenesis. Semin Cell Dev Biol 14: 151–157.

    Article  CAS  PubMed  Google Scholar 

  • Deftos ML, He YW, Ojala EW, Bevan MJ . (1998). Correlating notch signaling with thymocyte maturation. Immunity 9: 777–786.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Deftos ML, Huang E, Ojala EW, Forbush KA, Bevan MJ . (2000). Notch1 signaling promotes the maturation of CD4 and CD8 SP thymocytes. Immunity 13: 73–84.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Ellisen LW, Bird J, West DC, Soreng AL, Reynolds TC, Smith SD et al. (1991). TAN-1, the human homolog of the Drosophila notch gene, is broken by chromosomal translocations in T lymphoblastic neoplasms. Cell 66: 649–661.

    Article  CAS  PubMed  Google Scholar 

  • Fehling HJ, Krotkova A, Saint-Ruf C, von Boehmer H . (1995). Crucial role of the pre-T-cell receptor alpha gene in development of alpha beta but not gamma delta T cells. Nature 375: 795–798.

    Article  CAS  PubMed  Google Scholar 

  • Felli MP, Vacca A, Calce A, Bellavia D, Campese AF, Grillo R et al. (2005). PKC theta mediates pre-TCR signaling and contributes to Notch3-induced T-cell leukemia. Oncogene 24: 992–1000.

    Article  CAS  PubMed  Google Scholar 

  • Gay DL, Ramon H, Oliver PM . (2008). Cbl- and Nedd4-family ubiquitin ligases: balancing tolerance and immunity. Immunol Res 42: 51–64.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Groettrup M, Baron A, Griffiths G, Palacios R, von Boehmer H . (1992). T cell receptor (TCR) beta chain homodimers on the surface of immature but not mature alpha, gamma, delta chain deficient T cell lines. EMBO J 11: 2735–2745.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Hehner SP, Li-Weber M, Giaisi M, Droge W, Krammer PH, Schmitz ML . (2000). Vav synergizes with protein kinase C theta to mediate IL-4 gene expression in response to CD28 costimulation in T cells. J Immunol 164: 3829–3836.

    Article  CAS  PubMed  Google Scholar 

  • Hubbard EJ, Wu G, Kitajewski J, Greenwald I . (1997). sel-10, a negative regulator of lin-12 activity in Caenorhabditis elegans, encodes a member of the CDC4 family of proteins. Genes Dev 11: 3182–3193.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Irving BA, Alt FW, Killeen N . (1998). Thymocyte development in the absence of pre-T cell receptor extracellular immunoglobulin domains. Science 280: 905–908.

    Article  CAS  PubMed  Google Scholar 

  • Jehn BM, Dittert I, Beyer S, von der Mark K, Bielke W . (2002). c-Cbl binding and ubiquitin-dependent lysosomal degradation of membrane-associated Notch1. J Biol Chem 277: 8033–8040.

    Article  CAS  PubMed  Google Scholar 

  • Joutel A, Monet M, Domenga V, Riant F, Tournier-Lasserve E . (2004). Pathogenic mutations associated with cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy differently affect Jagged1 binding and Notch3 activity via the RBP/JK signaling Pathway. Am J Hum Genet 74: 338–347.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Kassenbrock CK, Anderson SM . (2004). Regulation of ubiquitin protein ligase activity in c-Cbl by phosphorylation-induced conformational change and constitude activation by tryosine to glutamate point mutations. J Biol Chem 279: 28017–28027.

    Article  CAS  PubMed  Google Scholar 

  • Liu Y, Liu YC, Meller N, Giampa L, Elly C, Doyle M et al. (1999). Protein kinase C activation inhibits tyrosine phosphorylation of Cbl and its recruitment of Src homology 2 domain-containing proteins. J Immunol 162: 7095–7101.

    CAS  PubMed  Google Scholar 

  • Liu YC . (2004). Ubiquitin ligases and the immune response. Annu Rev Immunol 22: 81–127.

    Article  PubMed  Google Scholar 

  • Logeat F, Bessia C, Brou C, LeBail O, Jarriault S, Seidah NG et al. (1998). The Notch1 receptor is cleaved constitutively by a furin-like convertase. Proc Natl Acad Sci USA 95: 8108–8112.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Montixi C, Langlet C, Bernard AM, Thimonier J, Dubois C, Wurbel MA et al. (1998). Engagement of T cell receptor triggers its recruitment to low-density detergent-insoluble membrane domains. EMBO J 17: 5334–5348.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Oberg C, Li J, Pauley A, Wolf E, Gurney M, Lendahl U . (2001). The Notch intracellular domain is ubiquitinated and negatively regulated by the mammalian Sel-10 homolog. J Biol Chem 276: 35847–35853.

    Article  CAS  PubMed  Google Scholar 

  • Ota S, Hazeki K, Rao N, Lupher Jr ML, Andoniou CE, Druker B et al. (2000). The RING finger domain of Cbl is essential for negative regulation of the Syk tyrosine kinase. J Biol Chem 275: 414–422.

    Article  CAS  PubMed  Google Scholar 

  • Panigada M, Porcellini S, Barbier E, Hoeflinger S, Cazenave PA, Gu H et al. (2002). Constitutive endocytosis and degradation of the pre-T cell receptor. J Exp Med 195: 1585–1597.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Pear WS, Aster JC, Scott ML, Hasserjian RP, Soffer B, Sklar J et al. (1996). Exclusive development of T cell neoplasms in mice transplanted with bone marrow expressing activated Notch alleles. J Exp Med 183: 2283–2291.

    Article  CAS  PubMed  Google Scholar 

  • Peschard P, Park M . (2003). Escape from Cbl-mediated downregulation: a recurrent theme for oncogenic deregulation of receptor tyrosine kinases. Cancer Cell 3: 519–523.

    Article  CAS  PubMed  Google Scholar 

  • Qiu L, Joazeiro C, Fang N, Wang HY, Elly C, Altman Y et al. (2000). Recognition and ubiquitination of Notch by Itch, a hect-type E3 ubiquitin ligase. J Biol Chem 275: 35734–35737.

    Article  CAS  PubMed  Google Scholar 

  • Rothenberg EV . (2001). Notchless T cell maturation? Nat Immunol 2: 189–190.

    Article  CAS  PubMed  Google Scholar 

  • Saint-Ruf C, Panigada M, Azogui O, Debey P, von Boehmer H, Grassi F . (2000). Different initiation of pre-TCR and gammadeltaTCR signalling. Nature 406: 524–527.

    Article  CAS  PubMed  Google Scholar 

  • Schweisguth F . (1999). Dominant-negative mutation in the beta2 and beta6 proteasome subunit genes affect alternative cell fate decisions in the Drosophila sense organ lineage. Proc Natl Acad Sci USA 96: 11382–11386.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Spolski R, Miescher G, Erard F, Risser R, MacDonald HR, Mak TW . (1988). Regulation of expression of T cell gamma chain, L3T4 and Ly-2 messages in Abelson/Moloney virus-transformed T cell lines. Eur J Immunol 18: 295–300.

    Article  CAS  PubMed  Google Scholar 

  • Talora C, Cialfi S, Oliviero C, Palermo R, Pascucci M, Frati L et al. (2006). Cross talk among Notch3, pre-TCR, and Tal1 in T-cell development and leukemogenesis. Blood 107: 3313–3320.

    Article  CAS  PubMed  Google Scholar 

  • Villalba M, Coudronniere N, Deckert M, Teixeiro E, Mas P, Altman A . (2000). A novel functional interaction between Vav and PKCtheta is required for TCR-induced T cell activation. Immunity 12: 151–160.

    Article  CAS  PubMed  Google Scholar 

  • Weng AP, Ferrando AA, Lee W, Morris JPT, Silverman LB, Sanchez-Irizarry C et al. (2004). Activating mutations of NOTCH1 in human T cell acute lymphoblastic leukemia. Science 306: 269–271.

    Article  CAS  PubMed  Google Scholar 

  • Wu G, Lyapina S, Das I, Li J, Gurney M, Pauley A et al. (2001). SEL-10 is an inhibitor of notch signaling that targets notch for ubiquitin-mediated protein degradation. Mol Cell Biol 21: 7403–7415.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

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Acknowledgements

We thank Dr A Joutel for Notch3 antibodies and Dr R Bei for purification of mouse monoclonal pTα antibody, Dr F Grassi for c-Cbl plasmid and Dr M Bouché for PKCθ plasmids. This work was supported by the Italian Association for Cancer Research (AIRC), the Italian Ministry of University and Research (MIUR), PRIN Program, the Italian Ministry of Health and the European Union (Euro-Thymaide FP6 Integrated Project; LHSB-CT-2003-543410).

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  1. S Checquolo and R Palermo: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Experimental Medicine, University ‘Sapienza’, Rome, Italy

    S Checquolo, R Palermo, S Cialfi, G Ferrara, C Oliviero, C Talora, D Bellavia, A Giovenco, P Grazioli, L Frati, A Gulino & I Screpanti

  2. Neuromed Institute, Pozzilli, Italy

    L Frati & A Gulino

  3. Institute Pasteur-Foundation Cenci Bolognetti, University ‘Sapienza’, Rome, Italy

    I Screpanti

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  1. S Checquolo
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Correspondence to I Screpanti.

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Supplementary Information accompanies the paper on the Oncogene website (http://www.nature.com/onc)

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Checquolo, S., Palermo, R., Cialfi, S. et al. Differential subcellular localization regulates c-Cbl E3 ligase activity upon Notch3 protein in T-cell leukemia. Oncogene 29, 1463–1474 (2010). https://doi.org/10.1038/onc.2009.446

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