Notch1: lineage specifier, oncogene and tumor suppressor

Notch proteins constitute a family of evolutionarily conserved transmembrane receptors found in organisms as diverse as worms and humans. They are activated by two distinct and equally conserved cell surface ligands called Delta and Serrate. Mammals have four receptors (Notch1-4) and five ligands: Jagged1 and Jagged2 (homologues of Serrate) and Delta1, Delta3 and Delta4 (homolgues of Delta) which are expressed on many cell types of various organs. These are fascinating proteins as they are involved in many differentiation processes and lineage decisions in fetal and postnatal development, as well as being implicated in self-renewal of tissues in organs such as the skin, the gut and the hematopoietic system. Stem cell maintenance, binary cell fate decisions and initiation of differentiation are among the many key functions controlled by Notch signaling. Since many of those functions are deregulated during tumorigenesis it is likely that Notch signaling may be directly involved in cancer. In fact the first mammalian homologue of drosophila Notch was identified by Jeff Sklar and colleagues in the early nineties in patients with T cell acute lymphoblastic leukemia 1. A chromosomal translocation that juxtaposes the C-terminal region of human NOTCH1 to the T cell receptor locus leads to aberrant expression of the cytoplasmic domain of NOTCH1. This causes the development of T cell leukemia showing the oncogenic potential of NOTCH1. Since then multiple Notch receptors and ligands have been found to be aberrantly expressed in various human carcinomas leading to a model for Notch function in cancer based on the ability of Notch to maintain normal precursor cells in a proliferating and undifferentiated state (reviewed in 2).

We are studying the role of Notch receptor and ligand family members in self-renewing organs such as the hematopoietic system and the skin by means of conditional gene targeting in the mouse. One particular goal is to compare the function of these gene family members in epithelial (skin) versus non-epithelial (hematopoietic) systems with respect to maintenance of stem cells, induction of differentiation and tumorigenesis. We have recently used the Cre-loxP recombination system to generate mice in which the Notch1 gene can be either conditionally inactivated in the bone marrow (BM) or at different developmental stages of thymocyte development. Using this system we showed that the Notch1 receptor is essential for T cell development, and furthermore is involved in T versus B cell fate determination. Notch1 deficient bone marrow progenitors migrate from the bone marrow to the thymus but can no longer be instructed towards the T cell fate by thymic epithelial cells expressing Notch ligands 3 4. Since these progenitors are still at least bi-potential they develop into B cells by default. At this particular level Notch1 acts as a lineage specifier.

In addition, we have been able to show that Notch1 signaling is involved in regulating V to DJβ-rearrangement 5 while it is dispensable at later stages of T cell development6. The molecular mechanisms and downstream targets by which Notch1 signaling exerts its essential functions during T cell development are currently under investigation.

In order to investigate whether Notch1 signaling also functions as a lineage specifier in epithelial tissues we have investigated its function in the skin. Notch1 signaling in murine keratinocytes stimulates expression of early differentiation markers such as keratin1 and involucrin, while late differentiation markers such as loricrin and Fillagrin are partially repressed. In addition Notch1 signaling induces expression of the CDKI p21 gene, which causes cell cycle arrest when cells leave the basal cell compartment to undergo terminal differentiation 7. In contrast to the previously established role of Notch1 as an oncogene in the hematopoietic system, we have identified Notch1 as a tumor suppressor in mouse skin. Inducible skin specific inactivation of Notch1 leads first to epidermal hyperprolieration and corneal hyperplasia, followed by the development of skin tumors. Notch1 deficiency in the epidermis results in sustained expression of Gli2 (a downstream effector of sonic hedgehog signaling), downregulation of the cell cycle regulator CDKI p21 and de-repressed β-catenin signaling in cells that should normally undergo differentiation 8.

Notch has multiple facets, one that specifies cell fates, one that promotes tumorigenisis and another that suppresses tumorigenisis. Which of these facets is shown is dependent on the cellular context and cross talk with other signal transduction pathways. It will be fascinating to uncover the mechanisms which lead to these apparently contradictory functions of Notch signaling. Only if we understand under which circumstances Notch functions as an oncogene or as a tumor suppressor, will we be able to come up with strategies to interfere with this pathway for therapeutic purposes.

Freddy Radtke
Ludwig Institute for Cancer Research, Lausanne Branch

 

Figure 1. Chromosomal translocation of NOTCH1 causes T cell leukaemia. The t(7:9) chromosomal translocation in T-ALL patients is characterised by the juxtaposition of the 3’ part of the human Notch1 gene into the T cell receptor β locus. This leads to expression of truncated Notch1 transcripts (akin to Notch1-IC) from the TCR? promoter, causing T cell leukaemia (T-ALL).

Figure 2. Notch1 function during T cell commitment and development. Hematopoietic stem cells (HSCs) give rise to an early lymphoid precursor (ELP) that makes a first cell fate decision producing either pro B cells or pro T cells. Subsequently pro T cells have to make a second lineage decision - either they commit to the γδ or to the αβ T cell lineage. Finally αβ committed thymocytes (at the DP (CD4+CD8+) stage) must choose between the CD4+ (CD4+CD8-) or the CD8+ (CD4-CD8+) T cell fate. Although multiple roles for Notch signaling during different lineage commitment processes have been postulated, the most decisive role for Notch1 signaling is its inductive role in the T/B lineage decision and its role during early intra-thymic T cell development by regulating V to DJβ rearrangements.

Figure 3. Notch1 function in mouse skin. Representation of mammalian skin with multiple layers of differentiating cells and proteins that are expressed in specific cellular layers (in green). Upon receiving differentiation signals, Notch1 induces expression of involucrin, Keratin1 (K1) and the cell cycle inhibitor Cip1, while preventing the onset of fillagrin and loricrin expression until the cell is at later stages of differentiation. Both β-catenin-mediated Wnt signaling and Gli2-mediated sonic hedgehog signaling are normally repressed in murine epidermis by Notch1. In Notch1-ablated skin, re-activation of the Wnt and sonic hedgehog pathways result in the development of basal-cell-carcinoma-like tumours in the mouse.

Radtke's Lab

 

References

  1. Ellisen L.W., Bird J., West D.C., Soreng A.L., Reynolds T.C., Smith S.D, Sklar J. TAN-1, the human homolog of the Drosophila notch gene, is broken by chromosomal translocations in T lymphoblastic neoplasms. Cell 66, 649-61. (1991).
  2. Radtke F. and Raj K. The role of Notch in tumorigenesis: oncogene or tumour suppressor? Nat Rev Cancer 10, 756-767 (2003).
  3. Radtke F., Wilson A., Stark G., Bauer M., van Meerwijk J., MacDonald H.R., Aguet M. Deficient T cell fate specification in mice with an induced inactivation of Notch1. Immunity 10, 547-558 (1999).
  4. Wilson A., MacDonald H.R. and Radtke F. Notch 1-deficient common lymphoid precursors adopt a B cell fate in the thymus. J Exp Med 194, 1003-12. (2001).
  5. Wolfer, A., Wilson, A., Nemir, M., MacDonald, H.R. and Radtke, F. Inactivation of Notch1 impairs VDJbeta rearrangement and allows pre-TCR-independent survival of early alpha beta Lineage Thymocytes. Immunity 16, 869-79. (2002).
  6. Wolfer A., Bakker T., Wilson A., Nicolas M., Ioannidis V., Littman D.R., Lee P.P., Wilson C.B., Held W., MacDonald H.R., Radtke F. Inactivation of Notch 1 in immature thymocytes does not perturb CD4 or CD8 T cell development. Nat Immunol 2, 235-41 (2001).
  7. Rangarajan A., Talora C., Okuyama R., Nicolas M., Mammucari C., Oh H., Aster J.C., Krishna S., Metzger D., Chambon P., Miele L., Aguet M., Radtke F., Dotto G.P. Notch signaling is a direct determinant of keratinocyte growth arrest and entry into differentiation. EMBO J 20, 3427-36. (2001).
  8. Nicolas, M., Wolfer A., Raj K., Kummer J.A., Mill P., van Noort H., Hui C.C., Clevers H., Dotto G.P., Radtke F. Notch1 functions as a tumor suppressor in mouse skin. Nat Genet 33, 416-21 (2003).