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Introduction Multiple factors control the size and composition of lymphocyte pools in secondary lymphoid organs. Both de novo production and attrition of mature lymphocytes through cell death or differentiation are [...], In lymphopenic environments, secondary lymphoid organs regulate the size of B and T cell compartments by supporting the homeostatic proliferation of mature lymphocytes. The molecular mechanisms underlying these responses and their functional consequences remain incompletely understood. To evaluate homeostasis of the mature B cell pool during lymphopenia, we turned to an adoptive transfer model of purified follicular B cells into [Rag2.sup.-/-] mouse recipients. Highly purified follicular B cells transdifferentiated into marginal zone-like B cells when transferred into [Rag2.sup.-/-] lymphopenic hosts but not into wild-type hosts. In lymphopenic spleens, transferred B cells gradually lost their follicular phenotype and acquired characteristics of marginal zone B cells, as judged by cell surface phenotype, expression of integrins and chemokine receptors, positioning close to the marginal sinus, and an ability to rapidly generate functional plasma cells. Initiation of follicular to marginal zone B cell transdifferentiation preceded proliferation. Furthermore, the transdifferentiation process was dependent on Notch2 receptors in B cells and expression of Delta-like 1 Notch ligands by splenic Ccl19-[Cre.sup.+] fibroblastic stromal cells. Gene expression analysis showed rapid induction of Notch-regulated transcripts followed by upregulated Myc expression and acquisition of broad transcriptional features of marginal zone B cells. Thus, naive mature B cells are endowed with plastic transdifferentiation potential in response to increased stromal Notch ligand availability during lymphopenia.

Introduction T cell acute lymphoblastic lymphoma (T-ALL) are serious hematologic malignancies of children and young adults. Current treatments that include intensive chemotherapy and cranial radiation are unsatisfactory, as they frequently [...], Somatic activating mutations in Notch1 contribute to the pathogenesis of T cell acute lymphoblastic lymphoma (T-ALL), but how activated Notch1 signaling exerts this oncogenic effect is not completely understood. Here we identify HIV-1 Rev-binding protein (Hrb), a component of the clathrin-mediated endocytosis machinery, as a critical mediator of Notch-induced T-ALL development in mice. Hrb was found to be a direct transcriptional target of Notch1, and Hrb loss reduced the incidence or delayed the onset of T-ALL in mouse models in which activated Notch1 signaling either contributes to or drives leukemogenesis. Consistent with this observation, Hrb supported survival and proliferation of hematopoietic and T cell precursor cells in vitro. We demonstrated that Hrb accelerated the uptake of transferrin, which was required for upregulation of the T cell protooncogene p21. Indeed, iron-deficient mice developed Notch1-induced T-ALL substantially more slowly than control mice, further supporting a critical role for iron uptake during leukemogenesis. Taken together, these results reveal that Hrb is a critical Notch target gene that mediates lymphoblast transformation and disease progression via its ability to satisfy the enhanced demands of transformed lympho-blasts for iron. Further, our data suggest that Hrb may be targeted to improve current treatment or design novel therapies for human T-ALL patients.

Introduction Phosphatase and tensin homolog deleted on chromosome 10 (PTEN) was first functionally characterized as a lipid phosphatase that catalyzes the reverse reaction of PI3K (1), (2). The major substrate [...], Mutations in the tumor-suppressor gene phosphatase and tensin homolog deleted on chromosome 10 (Pten) are associated with multiple cancers in humans, including T cell malignancies. Targeted deletion of Pten in T cells induces both a disseminated 'mature phenotype' lymphoma and a lymphoproliferative autoimmune syndrome in mice. Here, we have shown that these two diseases are separable and mediated by T lineage cells of distinct developmental stages. Loss of PTEN was found to be a powerful driver of lymphomagenesis within the thymus characterized by overexpression of the c-myc oncogene. In an otherwise normal thymic environment, PTEN-deficient T cell lymphomas invariably harbored RAG-dependent reciprocal t(14:15) chromosomal translocations involving the T cell receptor alpha/delta locus and c-myc, and their survival and growth was TCR dependent, but Notch independent. However, lymphomas occurred even if TCR recombination was prevented, although these lymphomas were less mature, arose later in life, and, importantly, were dependent upon Notch pathways to upregulate c-myc expression. In contrast, using the complementary methods of early thymectomy and adoptive transfers, we found that PTEN-deficient mature T cells were unable to undergo malignant transformation but were sufficient for the development of autoimmunity. These data suggest multiple and distinct regulatory roles for PTEN in the molecular pathogenesis of lymphoma and autoimmunity.

Tuberous sclerosis complex (TSC) is a dominantly inherited multisystem disease that is characterized by the formation of benign tumors in multiple organs (1), (2). The clinical manifestations of TSC are [...], Tuberous sclerosis complex (TSC) is a dominantly inherited disease that is characterized by the growth of multiple benign tumors that are often difficult to treat. TSC is caused by mutations that inactivate the TSC1 or TSC2 genes, which normally function to inhibit activation of mammalian target of rapamycin signaling. In this issue of the JCI, two studies reported by Karbowniczek et al. and Ma et al. link TSC inactivation with activated Notch signaling (see the related articles beginning on pages 93 and 103, respectively). Using a variety of approaches, both studies show that inactivation of TSC leads to Notch1 activation. Furthermore, studies in tumor cells suggest that inhibiting Notch slows growth of the tumor cells. Although much remains to be learned about the precise mechanisms by which TSC loss leads to Notch activation, the newly identified link of TSC to Notch provides the rationale for testing Notch inhibitors in TSC-associated tumors.

Introduction Development of the outflow tract (OFT) of the heart is complex, involving input from several embryologically distinct cell populations including cardiac progenitors from the second heart field, cardiac neural [...], Notch signaling is vital for proper cardiovascular development and function in both humans and animal models. Indeed, mutations in either JAGGED or NOTCH cause congenital heart disease in humans and NOTCH mutations are associated with adult valvular disease. Notch typically functions to mediate developmental interactions between adjacent tissues. Here we show that either absence of the Notch ligand Jagged1 or inhibition of Notch signaling in second heart field tissues results in murine aortic arch artery and cardiac anomalies. In mid-gestation, these mutants displayed decreased Fgf8 and Bmp4 expression. Notch inhibition within the second heart field affected the development of neighboring tissues. For example, faulty migration of cardiac neural crest cells and defective endothelial-mesenchymal transition within the outflow tract endocardial cushions were observed. Furthermore, exogenous Fgf8 was sufficient to rescue the defect in endothelial-mesenchymal transition in explant assays of endocardial cushions following Notch inhibition within second heart field derivatives. These data support a model that relates second heart field, neural crest, and endocardial cushion development and suggests that perturbed Notch-Jagged signaling within second heart field progenitors accounts for some forms of congenital and adult cardiac disease.

Introduction Notch1 belongs to a unique family of type I transmembrane receptors that regulate differentiation, proliferation, and survival in a dose- and context-dependent fashion (reviewed in ref. (1)). During transit [...], Gain-of-function NOTCH1 mutations are found in 50%-70% of human T cell acute lymphoblastic leukemia/lymphoma (T-ALL) cases. Gain-of-function NOTCH1 alleles that initiate strong downstream signals induce leukemia in mice, but it is unknown whether the gain-of-function NOTCH1 mutations most commonly found in individuals with T-ALL generate downstream signals of sufficient strength to induce leukemia. We addressed this question by expressing human gain-of-function NOTCH1 alleles of varying strength in mouse hematopoietic precursors. Uncommon gain-of-function NOTCH1 alleles that initiated strong downstream signals drove ectopic T cell development and induced leukemia efficiently. In contrast, although gain-of-function alleles that initiated only weak downstream signals also induced ectopic T cell development, these more common alleles failed to efficiently initiate leukemia development. However, weak gain-of-function NOTCH1 alleles accelerated the onset of leukemia initiated by constitutively active K-ras and gave rise to tumors that were sensitive to Notch signaling pathway inhibition. These data show that induction of leukemia requires doses of Notch1 greater than those needed for T cell development and that most NOTCH1 mutations found in T-ALL cells do not generate signals of sufficient strength to initiate leukemia development. Furthermore, low, nonleukemogenic levels of Notch1 can complement other leukemogenic events, such as activation of K-ras. Even when Notch1 participates secondarily, the resulting tumors show 'addiction' to Notch, providing a further rationale for evaluating Notch signaling pathway inhibitors in leukemia.

The cardiac outflow tract develops as a result of a complex interplay among several cell types, including cardiac neural crest cells, endothelial cells, and cardiomyocytes. In both humans and mice, [...]