Constitutive activation of STAT5 and Bcl-xL overexpression can induce endogenous erythroid colony formation in human primary cells.

The biologic hallmark of polycythemia vera (PV) is the formation of endogenous erythroid colonies (EECs) with an erythropoietin-independent differentiation. Recently, it has been shown that an activating mutation of JAK2 (V617F) was at the origin of PV. In this work, we studied whether the STAT5/Bcl-xL pathway could be responsible for EEC formation.

Apoptosis regulation in tetraploid cancer cells.

Tetraploidy can result in cancer-associated aneuploidy. As shown here, freshly generated tetraploid cells arising due to mitotic slippage or failed cytokinesis are prone to undergo Bax-dependent mitochondrial membrane permeabilization and subsequent apoptosis. Knockout of Bax or overexpression of Bcl-2 facilitated the survival of tetraploid cells at least as efficiently as the p53 or p21 knockout.

JAK2V617F expression in murine hematopoietic cells leads to MPD mimicking human PV with secondary myelofibrosis.

A JAK2(V617F) mutation is frequently found in several BCR/ABL-negative myeloproliferative disorders. To address the contribution of this mutant to the pathogenesis of these different myeloproliferative disorders, we used an adoptive transfer of marrow cells transduced with a retrovirus expressing JAK2(V617F) in recipient irradiated mice. Hosts were analyzed during the 6 months after transplantation.

Genetic and clinical implications of the Val617Phe JAK2 mutation in 72 families with myeloproliferative disorders.

To study the prevalence of the Val617Phe JAK2 mutation in familial cases of myeloproliferative disorder (MPD) and its possible implication as a predisposing genetic factor, we analyzed 72 families including 174 patients (81 polycythemia vera [PV], 68 essential thrombocythemia [ET], 11 myelofibrosis with myeloid metaplasia [MMM], 12 chronic myeloid leukemia), 1 systemic mastocytosis, and 1 chronic myelomonocytic leukemia (CMML). The JAK2 mutation was found in three quarters of patients with PV and MMM and in half of patients with ET. Among 46 families with at least 2 cases of PV, ET, or MMM, the JAK2 mutation was absent in 6 families, heterogeneously distributed in 18, and present in all MPD patients in 22.

Deficiency in the Wiskott-Aldrich protein induces premature proplatelet formation and platelet production in the bone marrow compartment.

The pathophysiology of microthrombocytopenia in the Wiskott-Aldrich syndrome (WAS) and its milder form, X-linked thrombocytopenia (XLT), is unclear. Although quantitative defects are correctable by splenectomy, residual platelet abnormalities are suggestive of intrinsic disturbances of production. In contrast to human patients, murine models of WASp deficiency exhibit only mild thrombocytopenia, and platelets are of normal size.

The SCL relative LYL-1 is required for fetal and adult hematopoietic stem cell function and B-cell differentiation.

Hematopoietic stem cells (HSCs) arise, self-renew, or give rise to all hematopoietic lineages through the effects of transcription factors activated by signaling cascades. Lyl-1 encodes a transcription factor containing a basic helix-hoop-helix (bHLH) motif closely related to scl/tal, which controls numerous decisions in embryonic and adult hematopoiesis. We report here that Lyl-1 null mice are viable and display normal blood cell counts, except for a reduced number of B cells resulting from a partial block after the pro-B stage.

A partial down-regulation of WASP is sufficient to inhibit podosome formation in dendritic cells.

The Wiskott Aldrich syndrome protein (WASP) is a hematopoietic-specific cytoskeletal regulator that is necessary for induction of normal immunity. In the context of effective gene therapy for WAS, cellular models of human WASP deficiency are important for definition of the threshold of protein expression required for optimal activity. Using lentiviral vector-mediated RNA interference (RNAi), we were able to down-regulate the levels of human WASP in cell lines and primary cells.

Kit-activating mutations cooperate with Spi-1/PU.1 overexpression to promote tumorigenic progression during erythroleukemia in mice.

The erythroleukemia developed by spi-1/PU.1 transgenic mice is a multistage process characterized by an early arrest of the proerythroblast differentiation followed later on by malignant transformation. Herein, we report the presence of acquired mutations in the SCF receptor gene (Kit) in 86% of tumors isolated during the late stage of the disease.

A unique activating mutation in JAK2 (V617F) is at the origin of polycythemia vera and allows a new classification of myeloproliferative diseases.

Myeloproliferative disorders (MPDs) are heterogeneous diseases that occur at the level of a multipotent hematopoietic stem cell. They are characterized by increased blood cell production related to cytokine hypersensitivity and virtually normal cell maturation. The molecular pathogenesis of the MPDs has been poorly understood, except for chronic myeloid leukemia (CML), where the Bcr-Abl fusion protein exhibits constitutive kinase activity.

Reduced retention of radioprotective hematopoietic cells within the bone marrow microenvironment in CXCR4-/- chimeric mice.

The physiologic role of CXCR4 on hematopoietic stem/progenitor cells (HSPCs) is not fully understood. Here, we show that radioprotection of lethally irradiated mice by embryonic day 14.5 (E14.

Mammalian target of rapamycin (mTOR) regulates both proliferation of megakaryocyte progenitors and late stages of megakaryocyte differentiation.

A major determinant in platelet production is the megakaryocyte (MK) size that is regulated both by ploidization and the increase in cytoplasmic volume at the end of maturation. Here we investigated the involvement of the mammalian target of rapamycin (mTOR) pathway in the regulation of megakaryopoiesis. We show that phosphorylation of mTOR, p70S6K1, and 4E-BP1 was diminished in thrombopoietin-cultured human MKs after rapamycin treatment.

A JAK2 mutation in myeloproliferative disorders: pathogenesis and therapeutic and scientific prospects.

Myeloproliferative disorders include several pathologies sharing the common feature of being clonal hematopoietic stem cell diseases. The molecular basis of chronic myeloid leukemia was characterized many years ago with the discovery of the t(9;22) translocation and its product the BCR-ABL oncoprotein. The recent finding of a recurrent mutation in the Janus 2 tyrosine kinase gene is a major advance in our understanding of the pathogenesis of several other myeloproliferative disorders, including polycythemia vera, essential thrombocythemia and idiopathic myelofibrosis.

An amphipathic motif at the transmembrane-cytoplasmic junction prevents autonomous activation of the thrombopoietin receptor.

Ligand binding to the thrombopoietin receptor (TpoR) is thought to impose a dimeric receptor conformation(s) leading to hematopoietic stem cell renewal, megakaryocyte differentiation, and platelet formation. Unlike other cytokine receptors, such as the erythropoietin receptor, TpoR contains an amphipathic KWQFP motif at the junction between the transmembrane (TM) and cytoplasmic domains. We show here that a mutant TpoR (delta5TpoR), where this sequence was deleted, is constitutively active.

JAK1 and Tyk2 activation by the homologous polycythemia vera JAK2 V617F mutation: cross-talk with IGF1 receptor.

The majority of polycythemia vera (PV) patients harbor a unique somatic mutation (V617F) in the pseudokinase domain of JAK2, which leads to constitutive signaling. Here we show that the homologous mutations in JAK1 (V658F) and in Tyk2 (V678F) lead to constitutive activation of these kinases. Their expression induces autonomous growth of cytokine-dependent cells and constitutive activation of STAT5, STAT3, mitogen-activated protein kinase, and Akt signaling in Ba/F3 cells.

Role of tyrosine kinases and phosphatases in polycythemia vera.

Protein tyrosine kinases (PTKs) and phosphatases (PTPs) play a crucial role in normal cell development, and dysfunction of these enzymes has been implicated in human cancers. Polycythemia vera (PV) is a clonal hematologic disease characterized by hypersensitivity of hematopoietic progenitor cells to growth factors and cytokines. Recently, a unique and clonal mutation in the JAK homology 2 (JH2) domain of JAK2 that results in a valine to phenylalanine substitution at position 617 (V617F) was found in the majority of PV patients.

Microarray analysis of LIF/Stat3 transcriptional targets in embryonic stem cells.

Mouse embryonic stem (ES) cells can be propagated in vitro while retaining their properties of pluripotency and self-renewal under the continuous presence of leukemia inhibitor factor (LIF). An essential role has been attributed to subsequent activation of the Stat3 transcription factor in mediating LIF self-renewal response. To date, however, downstream target genes of Stat3 in ES cells are still unknown.

RGS16 is a negative regulator of SDF-1-CXCR4 signaling in megakaryocytes.

Regulators of G-protein signaling (RGS) constitute a family of proteins involved in the negative regulation of signaling through heterotrimeric G protein-coupled receptors (GPCRs). Several RGS proteins have been implicated in the down-regulation of chemokine signaling in hematopoietic cells. The chemokine stromal-cell-derived factor 1 (SDF-1) activates migration of hematopoietic progenitors cells but fails to activate mature megakaryocytes despite high levels of CXC chemokine receptor 4 (CXCR4) receptor expression in these cells.

Osteopontin is upregulated by BCR-ABL.

Chronic myelogenous leukemia (CML) is characterized by its hallmark oncogene BCR-ABL and the progression from a chronic phase toward an acute leukemia, with a differentiation arrest of the leukemic clone. In the present study, we conducted a microarray analysis using an inducible model of BCR-ABL expression based on the TET-OFF system, and we found that osteopontin (OPN), a component of stem cell niche, is overexpressed in BCR-ABL-expressing cells. Studies using mutant forms of BCR-ABL demonstrated that the BCR-ABL-induced OPN overexpression was a tyrosine kinase-dependent event.

Recurrent V75M mutation within the Wiskott-Aldrich syndrome protein: description of a homozygous female patient.

The Wiskott-Aldrich syndrome is a rare genetic disorder due to mutations in the WAS gene situated on chromosome X. It is comprised of microthrombocytopenia, eczema and immunodeficiency. However, the phenotypical presentation may vary as to the number and intensity of its manifestations.

[Hope and limits in cell therapies].

The hematopoietic system is one of the best characterized human cellular system in which a multipotent adult stem cell is at the origin of all the cells of a tissue. These last years, the use of stem cells has given rise to many hopes in regenerative medicine, especially in diseases without efficient therapies. However the hematopoietic system in which stem cell transplantation has entered in clinical practice for many years has also shown the limits of these approaches.

Overexpression of sphingosine kinase 1 is an oncogenic event in erythroleukemic progression.

The erythroleukemia developed by spi-1/PU.1-transgenic mice is a model of multistage oncogenic process. Isolation of tumor cells representing discrete stages of leukemic progression enables the dissection of some of the critical events required for malignant transformation.

Role for the nuclear factor kappaB pathway in transforming growth factor-beta1 production in idiopathic myelofibrosis: possible relationship with FK506 binding protein 51 overexpression.

The release of transforming growth factor-beta1 (TGF-beta1) in the bone marrow microenvironment is one of the main mechanisms leading to myelofibrosis in murine models and probably in the human idiopathic myelofibrosis (IMF). The regulation of TGF-beta1 synthesis is poorly known but seems regulated by nuclear factor kappaB (NF-kappaB). We previously described the overexpression of an immunophilin, FK506 binding protein 51 (FKBP51), in IMF megakaryocytes.

p210BCR-ABL inhibits SDF-1 chemotactic response via alteration of CXCR4 signaling and down-regulation of CXCR4 expression.

It has been shown that p210(BCR-ABL) significantly impairs CXCR4 signaling. We report here that the migratory response to SDF-1 was profoundly altered in blast crisis, whereas chronic-phase CD34(+) cells migrated normally to this chemokine. This migratory defect was associated with a low CXCR4 membrane expression.