The AML1-CBF beta transcription factor is the most frequent target of chromosomal rearrangements in human leukemia. To investigate its normal function, we generated mice lacking AML1. Embryos with homozygous mutations in AML1 showed normal morphogenesis and yolk sac-derived erythropoiesis, but lacked fetal liver hematopoiesis and died around E12.5. Sequentially targeted AML1-/-es cell retained their capacity to differentiate into primitive erythroid cells in vitro; however, no myeloid or erythroid progenitors of definitive hematopoietic origin were detected in either the yolk sac or fetal livers of mutant embryos. Moreover, this hematopoietic defect was intrinsic to the stem cells in that AML1-/-ES cells failed to contribute to hematopoiesis in chimeric animals. These results suggest that AML1-regulated target genes are essential for definitive hematopoiesis of all lineages.

Mouse CD4+CD8+ double-positive (DP) thymocytes differentiate into CD4+ helper-lineage cells upon expression of the transcription factor Th-POK but commit to the CD8+ cytotoxic lineage in its absence. We report the redirected differentiation of class I-restricted thymocytes into CD4+CD8- helper-like T cells upon loss of Runx transcription factor complexes. A Runx-binding sequence within the Th-POK locus acts as a transcriptional silencer that is essential for Th-POK repression and for development of CD8+ T cells. Thus, Th-POK expression and genetic programming for T helper cell development are actively inhibited by Runx-dependent silencer activity, allowing for cytotoxic T cell differentiation. Identification of the transcription factors network in CD4 and CD8 lineage choice provides insight into how distinct T cell subsets are developed for regulating the adaptive immune system.

In this study, we examined a large number of patients to clarify FLT3 gene has some structural similarities including the the distribution and frequency of a recently described FLT3 tannumber of exons, size of exons and exon/intron boundaries dem duplication among hematopoietic malignancies, including with genes RTKs, FMS, KIT, and platelet-derived growth-factor of 24 showed internal tandem duplication with or withoutWe recently demonstrated internal tandem duplication insertion of nucleotides. In one AML, insertion and deletion within JM/TK-I domains as a somatic mutation of FLT3 found without duplication was determined. All 24 lengthened in 17% of patients with acute myelogenous leukemia (AML). 14 sequences were in-frame. Duplication takes place in the Since this mutation was not found in any patients with acute sequence coding for the JM domain and leaves the TK domain lymphocytic leukemia, we described that this mutation could intact. In conclusion, we emphasize that the length mutation of FLT3 at JM/TK-I domains were restricted to AML and MDS.be specific in myeloid malignancies. To clarify the incidence Since all these mutations resulted in in-frame, this abnormality and distribution of the FLT3 mutation among hematological might function for the proliferation of leukemic cells.malignancies, we examined a large number of patients with

Mrg class G-protein-coupled receptors (GPCRs) are expressed exclusively in sensory neurons in the trigeminal and dorsal root ganglia. Pharmacological activation of Mrg proteins is capable of modulating sensory neuron activities and elicits nociceptive effects. In this study, we illustrate a control mechanism that allows the Runx1 runt domain transcription factor to generate compartmentalized expression of these sensory GPCRs. Expression of MrgA, MrgB, and MrgC subclasses is confined to an "A/B/C" neuronal compartment that expresses Runx1 transiently (or does not express Runx1), whereas MrgD expression is restricted to a "D" compartment with persistent Runx1 expression.

It has been well established that a number of transcription factors play critical roles in regulating the fate of hematopoietic stem cell populations. One of them is the leukemia-associated transcription factor acute myeloid leukemia 1 (AML1; also known as runt-related transcription factor 1, or RUNX1). This gene was originally cloned from the breakpoint of the t(8;21) reciprocal chromosome translocation and was later recognized as one of the most frequent targets of leukemia-associated gene aberrations. Gene-targeting experiments revealed that transcriptionally active AML1 is essential for the establishment of definitive hematopoiesis. More specifically, this gene functions in the emergence of the hematopoietic progenitor cells from the hemogenic endothelium by budding in the aorta-gonad-mesonephros region, and its expression points to the sites with strong potential for the emergence of hematopoietic stem cells. This review discusses aspects of the biologic properties of AML1 in early hematopoietic development.

The t(8;21)-encoded AML1-ETO chimeric product is believed to be causally involved in up to 15% of acute myelogenous leukemias through an as yet unknown mechanism. To directly investigate the role of AML1-ETO in leukemogenesis, we used gene targeting to create anAML1-ETO “knock-in” allele that mimics the t(8;21). Unexpectedly, embryos heterozygous for AML1-ETO(AML1-ETO/+) died around E13.5 from a complete absence of normal fetal liver–derived definitive hematopoiesis and lethal hemorrhages. This phenotype was similar to that seen following homozygous disruption of either AML1 orCBFβ. However, in contrast to AML1- or CBFβ-deficient embryos, fetal livers from AML1-ETO/+ embryos contained dysplastic multilineage hematopoietic progenitors that had an abnormally high self-renewal capacity in vitro. To further document the role of AML1-ETO in these growth abnormalities, we used retroviral transduction to express AML1-ETO in murine adult bone marrow–derived hematopoietic progenitors. AML1-ETO–expressing cells were again found to have an increased self-renewal capacity and could be readily established into immortalized cell lines in vitro. Taken together, these studies suggest that AML1-ETO not only neutralizes the normal biologic activity of AML1 but also directly induces aberrant hematopoietic cell proliferation.

AML1 is one of the most frequently mutated genes associated with human acute leukemia and encodes the DNA-binding subunit of the heterodimering transcriptional factor complex, core-binding factor (CBF) (or polyoma enhancer binding protein 2 [PEBP2]). A null mutation in either AML1 or its dimerizing partner, CBF␤, results in embryonic lethality secondary to a complete block in fetal liver hematopoiesis, indicating an essential role of this transcription complex in the development of definitive hematopoiesis. The hematopoietic phenotype that results from the loss of AML1 can be replicated in vitro with a two-step culture system of murine embryonic stem (ES) cells. Using this experimental system, we now demonstrate that this hematopoietic defect can be rescued by expressing the PEBP2␣B1 (AML1b) isoform under the endogenous AML1-regulatory sequences through a knock-in (targeted insertion) approach. Moreover, we demonstrate that the rescued AML1؊/؊ ES cell clones contribute to lymphohematopoiesis within the context of chimeric animals. Rescue requires the transcription activation domain of AML1 but does not require the C-terminal VWRPY motif, which is conserved in all AML1 family members and has been shown to interact with the transcriptional corepressor, Groucho/transducin-like Enhancer of split. Taken together, these data provide compelling evidence that the phenotype seen in AML1-deficient mice is due solely to the loss of transcriptionally active AML1.Hematopoietic development in mammals is supported by two discrete cellular populations which are believed to be derived from distinct cellular origins (5, 22). In the mouse, for example, the first wave of primitive hematopoiesis emerges around day 7.5 postcoitus (E 7.5) in the yolk sac and consists primarily of large nucleated primitive erythrocytes, which diminish midgestation. By contrast, around E 9.5, the second wave of hematopoiesis emerges in the fetal liver and consists of so-called definitive hematopoietic cells, including enucleated erythrocytes containing adult-type globin molecules, myeloid cells, and lymphoid progenitors. The site of this second wave of hematopoiesis is subsequently shifted to the bone marrow and spleen prior to birth.Recent studies have revealed that a number of transcriptional factors play critical roles in regulating the fate of the hematopoietic stem cell populations. These factors are divided into two groups: one contains factors which regulate both primitive and definitive hematopoiesis, and the other contains factors whose activities are required for the development of some or all of the definitive hematopoietic lineages (reviewed in reference 44). The acute myeloid leukemia (AML) 1 gene, AML1, belongs to the latter group and is unique in that loss of this gene in mice results in the complete absence of all definitive hematopoietic lineages (37, 51).AML1 was originally cloned from the breakpoint of chromosome 21 in t(8;21)(q22;q22), which is associated with 40% of the AML cases of the French-British-American classification M2 subt...

AML1/Runx1 is a frequent target of leukemia-associated gene aberration, and it encodes a transcription factor essential for definitive hematopoiesis. We previously reported that the AML1 molecules with trans-activation subdomains retained can rescue in vitro hematopoietic defects of AML1-deficient mouse embryonic stem (ES) cells when expressed by using a knock-in approach. Extending this notion to in vivo conditions, we found that the knock-in ES cell clones with AML1 mutants, which retain trans-activation subdomains but lack C-terminal repression subdomains including the conserved VWRPY motif, contribute to hematopoietic tissues in chimera mice. We also found that germline mice homozygous for the mutated AML1 allele, which lacks the VWRPY motif, exhibit a minimal effect on hematopoietic development, as was observed in control knock-in mice with full-length AML1. On the other hand, reduced cell numbers and deviant CD4 expression were observed during early T-lymphoid ontogeny in the VWRPY-deficient mice, whereas the contribution to the thymus by the corresponding ES cell clones was inadequate. These findings demonstrate that AML1 with its trans-activating subdomains is essential and sufficient for hematopoietic development in the context of the entire mouse. In addition, its transrepression activity, depending on the Cterminal VWRPY motif, plays a role in early thymocyte development. IntroductionVertebrate hematopoietic development is characterized by the sequential appearance of two cell populations, further defined as primitive and definitive hematopoiesis. 1 In mice, for example, primitive hematopoiesis is first seen in the form of blood islands in the yolk sac of the 7.5-day-old (E7.5) mouse embryo. It is thought that this cell population is directly differentiated from hemangioblasts, a bipotent precursor. Primitive hematopoiesis consists predominantly of a large and nucleated erythroid population containing embryonic-type hemoglobin. In contrast to the restricted and temporal development of this first wave, which diminishes at midgestation, definitive hematopoiesis originates from the aorta-gonad-mesonephros (AGM) region, where stem cells with long-term repopulating ability of multilineage hematopoiesis emerge at approximately E9.5, as a result of budding, from the ventral endothelial cells of the great vessels. 2,3 The stem cells then migrate into the fetal liver and proliferate to rapidly establish the definitive hematopoiesis of all lineages, including progenitors for T-and B-lymphoid populations. Active sites for definitive hematopoiesis are transferred to bone marrow and spleen before birth and function throughout life within these organs.These stem cells are equipped with a number of critical transcription factors that play pivotal roles in determining the fate of the cells at discrete developmental stages. Most of these molecules have been identified by isolating DNA-binding proteins to known cis-regulatory elements of lineage-specific genes or by cloning the DNA targets of leukemia-associated chrom...