During progression of Moloney murine leukemia virus (Mo-MuLV)-induced rat T cell lymphomas, growth selection results in the expansion of cell clones carrying increasing numbers of integrated proviruses. These new provirus insertions reproducibly contribute to enhanced growth, allowing the emergence of cell clones from the initially heterogeneous population of tumor cells. The Mo-MuLV-induced rat T cell lymphoma lines 2780d and 5675d, which are dependent on interleukin-2 (IL-2) for growth in culture (IL-2d), were placed in IL-2-free medium to select for IL-2-independent (IL-2i) mutants. Southern blot analysis of genomic DNA from these mutants, which was hybridized to a Mo-MuLV long terminal repeat probe, revealed that all mutants carried new provirus insertions (from one to four new proviruses per cell line). A locus of integration identified through cloning of the single new provirus detected in one of the IL-2i mutants, 2780i.5, was found to be the target of provirus insertion in 1 additional IL-2i cell line of 24 tested. A full-length cDNA of a gene (growth factor independence-1 [Gfi-11) activated by promoter insertion in the 2780i.5 cells was cloned and shown to encode a novel zinc finger protein. Gfi-1 is expressed at low levels in IL-2d cell lines cultured in IL-2-containing medium and at high levels in most IL-2i cell lines, including the two harboring a provirus at this locus. Gfi-1 expression in adult animals is restricted to the thymus, spleen, and testis. In mitogen-stimulated splenocytes, Gfi-l expression begins to rise at 12 h after stimulation and reaches very high levels after 50 h, suggesting that it may be functionally involved in events occurring after the interaction of IL-2 with its receptor, perhaps during the transition from the G, to the S phase of the cell cycle. In agreement with this, Gfi-I does not induce the expression of IL-2. Expression of Gfi-1 in 2780d cells following transfer of a Gfi-1/LXSN retrovirus construct contributes to the emergence of the IL-2i phenotype.The interaction between interleukin-2 (IL-2) and the highaffinity interleukin-2 receptor (IL-2R) is a critical event in T cell activation, triggering proliferation of cells after antigen binding to the T cell receptor (1,8,42,47). This clonal proliferation is obligatory for immunocompetence; mice rendered IL-2 deficient by targeted disruption of the IL-2 gene show decreased proliferative responses to mitogen and decreased T helper cell activity (40), while humans without functional IL-2 have severe combined immunodeficiency (31, 49).The events prior to and after the interaction of IL-2 with its receptor have been explored to date by using several strategies, including the screening of T cells during activation for the expression of known genes (35), the use of subtraction cDNA libraries to clone genes whose expression is altered in activated T cells compared with in resting T cells (23), and the identification and cloning either of genes encoding proteins which interact with signalling molecules or of genes known to regul...

The Tpl-2 locus, doned by provirus tagging from one of three sublines of the Moloney leukemia virusinduced rat thymoma 2769, defines a gene encoding a protein kinase associated with progression in 22.5% of the tumors.

Gfi-1 is a cellular proto-oncogene that was identified as a target of provirus integration in T-cell lymphoma lines selected for interleukin-2 (IL-2) independence in culture and in primary retrovirus-induced lymphomas. Gfi-1 encodes a zinc finger protein that functions as a transcriptional repressor. Here we show that Gfi-1B, a Gfi-1 related gene expressed in bone marrow and spleen, also encodes a transcriptional repressor. IL-6-induced G 1 arrest and differentiation of the myelomonocytic cell line M1 were linked to the downregulation of Gfi-1B and the parallel induction of the cyclin-dependent kinase inhibitor p21WAF1 . Experiments addressing the potential mechanism of the apparent coordinate regulation of these genes revealed that Gfi-1B represses p21 WAF1 directly by binding to a high-affinity site at ؊1518 to ؊1530 in the p21 WAF1 promoter. Forced expression of Gfi-1B, but not of Gfi-1B deletion mutants lacking the repressor domain, blocked the IL-6-mediated induction of p21 WAF1 and inhibited G 1 arrest and differentiation. We conclude that Gfi-1B is a direct repressor of the p21 WAF1 promoter, the first such repressor identified to date, and that sustained expression of Gfi-1B blocks IL-6-induced G 1 arrest and differentiation of M1 cells perhaps because it prevents p21 WAF1induction by IL-6.Hematopoiesis is a process that takes place in the bone marrow throughout the life of an individual. During this process a small number of hematopoietic stem cells respond to microenvironmental cues to either divide and self-renew or differentiate into hematopoietic progenitors committed to specic hematopoietic lineages. The committed hematopoietic progenitors, in turn, also undergo self-renewal or terminal differentiation. The maintenance of the hematopoietic stem cells and their selection, commitment, and maturation along different hematopoietic lineages depend on cell-to-cell and cell-tostroma interactions, secreted cytokines, and intracellular signaling molecules (45,46). The molecular mechanisms involved in regulating hematopoietic cell commitment and differentiation can be addressed in differentiating hematopoietic tissues in intact animals (64) and in cell lines that can be induced to differentiate (35,38). With both systems, a number of molecules, including growth factors, receptors, and transcription factors, have been identified and shown to contribute to hematopoiesis in a hierarchical order (10,29,42,65).The myelomonocytic cell line M1 undergoes G 1 arrest and differentiation following exposure to interleukin-6 (IL-6) or leukemia inhibitory factor (15, 50). During this process the expression of several signaling molecules is altered. c-myb is downregulated within 3 h from the start of the exposure to IL-6 or leukemia inhibitory factor (26). This is followed by the downregulation of c-myc (25). Overexpression of either c-myb or c-myc inhibits IL-6-induced differentiation of M1 cells (25,26,49), suggesting that the downregulation of these molecules is required for differentiation. Another molecule whose expre...

A common integration site, cloned from MoMuLV-induced rat T cell lymphomas, was mapped immediately upstream of Not dead yet-1 (Ndy1)/KDM2B, a gene expressed primarily in testis, spleen, and thymus, that is also known as FBXL10 or JHDM1B. Ndy1 encodes a nuclear, chromatin-associated protein that harbors Jumonji C (JmjC), CXXC, PHD, proline-rich, F-box, and leucine-rich repeat domains. Ndy1 and its homolog Ndy2/KDM2A (FBXL11 or JHDM1A), which is also a target of provirus integration in retrovirus-induced lymphomas, encode proteins that were recently shown to possess Jumonji C-dependent histone H3 K36 dimethyldemethylase or histone H3 K4 trimethyl-demethylase activities. Here, we show that mouse embryo fibroblasts engineered to express Ndy1 or Ndy2 undergo immortalization in the absence of replicative senescence via a JmjC domain-dependent process that targets the Rb and p53 pathways. Knockdown of endogenous Ndy1 or expression of JmjC domain mutants of Ndy1 promote senescence, suggesting that Ndy1 is a physiological inhibitor of senescence in dividing cells and that inhibition of senescence depends on histone H3 demethylation.cancer ͉ histone demethylase ͉ immortalization ͉ senescence ͉ insertional mutagenesis