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Koichi Akashi

Publications and source records attributed to Koichi Akashi.

At least 19 recordsLinked to original sources

Large-scale genome analysis of bovine commensal Escherichia coli reveals that bovine-adapted E. coli lineages are serving as evolutionary sources of the emergence of human intestinal pathogenic strains.

How pathogens evolve their virulence to humans in nature is a scientific issue of great medical and biological importance. Shiga toxin (Stx)-producing Escherichia coli (STEC) and enteropathogenic E. coli (EPEC) are the major foodborne pathogens that can cause hemolytic uremic syndrome and infantile diarrhea, respectively. The locus of enterocyte effacement (LEE)-encoded type 3 secretion system (T3SS) is the major virulence determinant of EPEC and is also possessed by major STEC lineages. Cattle are thought to be the primary reservoir of STEC and EPEC. However, genome sequences of bovine commensal E. coli are limited, and the emerging process of STEC and EPEC is largely unknown. Here, we performed a large-scale genomic comparison of bovine commensal E. coli with human commensal and clinical strains, including EPEC and STEC, at a global level. The analyses identified two distinct lineages, in which bovine and human commensal strains are enriched, respectively, and revealed that STEC and EPEC strains have emerged in multiple sublineages of the bovine-associated lineage. In addition to the bovine-associated lineage-specific genes, including fimbriae, capsule, and nutrition utilization genes, specific virulence gene communities have been accumulated in stx- and LEE-positive strains, respectively, with notable overlaps of community members. Functional associations of these genes probably confer benefits to these E. coli strains in inhabiting and/or adapting to the bovine intestinal environment and drive their evolution to highly virulent human pathogens under the bovine-adapted genetic background. Our data highlight the importance of large-scale genome sequencing of animal strains in the studies of zoonotic pathogens.

Animals↗

The order of expression of transcription factors directs hierarchical specification of hematopoietic lineages.

The mechanism of lineage specification in multipotent stem cells has not been fully understood. We recently isolated progenitors with the eosinophil, basophil, or mast cell lineage potential, all of which originate from granulocyte/monocyte progenitors (GMPs). By using these prospectively purified progenitors, we show here that the expression timing of GATA-2 and CCAAT enhancer-binding protein alpha (C/EBPalpha) can differentially control their lineage commitment. The expression of GATA-2 instructed C/EBPalpha-expressing GMPs to commit exclusively into the eosinophil lineage, while it induced basophil and/or mast cell lineage commitment if C/EBPalpha was suppressed at the GMP stage. Furthermore, simply by switching the order of C/EBPalpha and GATA-2 transduction, even lymphoid-committed progenitors recaptured these developmental processes to be reprogrammed into each of these lineages. We propose that the order of expression of key transcription factors is critical for their interplay to selectively drive lineage specification programs, by which stem cells could generate multiple lineage cells in a hierarchical manner.

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Essential role for cyclin D3 in granulocyte colony-stimulating factor-driven expansion of neutrophil granulocytes.

The proliferation of neutrophil granulocyte lineage is driven largely by granulocyte colony-stimulating factor (G-CSF) acting via the G-CSF receptors. In this study, we show that mice lacking cyclin D3, a component of the core cell cycle machinery, are refractory to stimulation by the G-CSF. Consequently, cyclin D3-null mice display deficient maturation of granulocytes in the bone marrow and have reduced levels of neutrophil granulocytes in their peripheral blood. The mutant mice are unable to mount a normal response to bacterial challenge and succumb to microbial infections. In contrast, the expansion of hematopoietic stem cells and lineage-committed myeloid progenitors proceeds relatively normally in mice lacking cyclin D3, revealing that the requirement for cyclin D3 function operates at later stages of neutrophil development. Importantly, we verified that this requirement is specific to cyclin D3, as mice lacking other G(1) cyclins (D1, D2, E1, or E2) display normal granulocyte counts. Our analyses revealed that in the bone marrow cells of wild-type mice, activation of the G-CSF receptor leads to upregulation of cyclin D3. Collectively, these results demonstrate that cyclin D3 is an essential cell cycle recipient of G-CSF signaling, and they provide a molecular link of how G-CSF-dependent signaling triggers cell proliferation.

Animals↗

C/EBPbeta is required for 'emergency' granulopoiesis.

During 'emergency' situations such as infections, host defense requires rapid mobilization of bone marrow granulocyte progenitors. 'Steady-state' granulopoiesis is absolutely dependent on the C/EBPalpha transcription factor, but the transcriptional mechanisms underlying emergency granulopoiesis remain unclear. Here we show that large numbers of granulocytes were generated from C/EBPalpha-deficient progenitors after cytokine stimulation in vivo. Cytokine treatment or fungal infection induced upregulation of C/EBPbeta but not C/EBPalpha or C/EBPepsilon transcripts in granulocyte progenitors, and C/EBPbeta-deficient progenitors showed decreased emergency-induced granulopoiesis in vitro and in vivo. C/EBPbeta inhibited proliferation less severely than did C/EBPalpha. These data suggest a critical function for C/EBPbeta in emergency granulopoiesis, which demands both differentiation and proliferation of granulocyte precursors.

Animals↗

Oncogenic K-ras cooperates with PML-RAR alpha to induce an acute promyelocytic leukemia-like disease.

Most patients with acute promyelocytic leukemia (APL) express PML-RAR alpha, the fusion product of t(15;17)(q22;q11.2). Transgenic mice expressing PML-RAR alpha develop APL with long latency, low penetrance, and acquired cytogenetic abnormalities. Based on observations that 4% to 10% of APL patients harbor oncogenic ras mutations, we coexpressed oncogenic K-ras from its endogenous promoter with PML-RAR alpha to generate a short-latency, highly penetrant mouse model of APL. The APL disease was characterized by splenomegaly, leukocytosis, extramedullary hematopoiesis (EMH) in spleen and liver with an increased proportion of immature myeloperoxidase-expressing myeloid forms; transplantability to secondary recipients; and lack of cytogenetic abnormalities. Bone marrow cells showed enhanced self-renewal in vitro. This model establishes a role for oncogenic ras in leukemia pathogenesis and thus validates the oncogenic RAS signaling pathway as a potential target for therapeutic inhibition in leukemia patients. This mouse model should be useful for investigating signaling pathways that promote self-renewal in APL and for testing the in vivo efficacy of RAS signaling pathway inhibitors in conjunction with other targeted therapies such as ATRA (all trans retinoic acid) and arsenic trioxide.

Animals↗

Thymus exclusivity: all the right conditions for T cells.

A fraction of primitive "lymphoid" precursors retain plasticity for myeloid differentiation. In this issue of Immunity, Laiosa et al. describe that Notch-Delta signals can protect thymic precursors from reprogramming into the myeloid lineage, antagonizing the enforced myeloid transcription factors such as PU.1 and C/EBPalpha.

Animals↗

Developmental checkpoints of the basophil/mast cell lineages in adult murine hematopoiesis.

Basophils and mast cells, which are selectively endowed with the high-affinity IgE receptor and mediate a range of adaptive and innate immune responses, have an unknown developmental relationship. Here, by evaluating the expression of the beta7 integrin, a molecule that is required for selective homing of mast cell progenitors (MCPs) to the periphery, we identified bipotent progenitors that are capable of differentiating into either cell type in the mouse spleen. These basophil/mast cell progenitors (BMCPs) gave rise to basophils and mast cells at the single-cell level and reconstituted both mucosal and connective tissue mast cells. We also identified the basophil progenitor (BaP) and the MCP in the bone marrow and the gastrointestinal mucosa, respectively. We further show that the granulocyte-related transcription factor CCAAT/enhancer-binding protein alpha (C/EBPalpha) plays a primary role in the fate decision of BMCPs, being expressed in BaPs but not in MCPs. Thus, circulating basophils and tissue mast cells share a common developmental stage at which their fate decision might be controlled principally by C/EBPalpha.

Animals↗

Lymphoid cell growth and transformation are suppressed by a key regulatory element of the gene encoding PU.1.

Tight regulation of transcription factors, such as PU.1, is crucial for generation of all hematopoietic lineages. We previously reported that mice with a deletion of an upstream regulatory element (URE) of the gene encoding PU.1 (Sfpi1) developed acute myeloid leukemia. Here we show that the URE has an essential role in orchestrating the dynamic PU.1 expression pattern required for lymphoid development and tumor suppression. URE deletion ablated B2 cells but stimulated growth of B1 cells in mice. The URE was a PU.1 enhancer in B cells but a repressor in T cell precursors. TCF transcription factors coordinated this repressor function and linked PU.1 to Wnt signaling. Failure of appropriate PU.1 repression in T cell progenitors with URE deletion disrupted differentiation and induced thymic transformation. Genome-wide DNA methylation assessment showed that epigenetic silencing of selective tumor suppressor genes completed PU.1-initiated transformation of lymphoid progenitors with URE deletion. These results elucidate how a single transcription factor, PU.1, through the cell context-specific activity of a key cis-regulatory element, affects the development of multiple cell lineages and can induce cancer.

Animals↗

Identification of eosinophil lineage-committed progenitors in the murine bone marrow.

Eosinophil lineage-committed progenitors (EoPs) are phenotypically isolatable in the steady-state murine bone marrow. Purified granulocyte/monocyte progenitors (GMPs) gave rise to eosinophils as well as neutrophils and monocytes at the single cell level. Within the short-term culture of GMPs, the eosinophil potential was found exclusively in cells activating the transgenic reporter for GATA-1, a transcription factor capable of instructing eosinophil lineage commitment. These GATA-1-activating cells possessed an IL-5Ralpha(+)CD34(+)c-Kit(lo) phenotype. Normal bone marrow cells also contained IL-5Ralpha(+)CD34(+)c-Kit(lo) EoPs that gave rise exclusively to eosinophils. EoPs significantly increased in number in response to helminth infection, suggesting that the EoP stage is physiologically involved in eosinophil production in vivo. EoPs expressed eosinophil-related genes, such as the eosinophil peroxidase and the major basic protein, but did not express basophil/mast cell-related mast cell proteases. The enforced retroviral expression of IL-5Ralpha in GMPs did not enhance the frequency of eosinophil lineage read-outs, whereas IL-5Ralpha(+) GMPs displayed normal neutrophil/monocyte differentiation in the presence of IL-5 alone. Thus, IL-5Ralpha might be expressed specifically at the EoP stage as a result of commitment into the eosinophil lineage. The newly identified EoPs could be the cellular target in the treatment of a variety of disorders mediated by eosinophils.

Animals↗

Development of functional human blood and immune systems in NOD/SCID/IL2 receptor {gamma} chain(null) mice.

Here we report that a new nonobese diabetic/severe combined immunodeficient (NOD/SCID) mouse line harboring a complete null mutation of the common cytokine receptor gamma chain (NOD/SCID/interleukin 2 receptor [IL2r] gamma(null)) efficiently supports development of functional human hemato-lymphopoiesis. Purified human (h) CD34(+) or hCD34(+)hCD38(-) cord blood (CB) cells were transplanted into NOD/SCID/IL2rgamma(null) newborns via a facial vein. In all recipients injected with 10(5) hCD34(+) or 2 x 10(4) hCD34(+)hCD38(-) CB cells, human hematopoietic cells were reconstituted at approximately 70% of chimerisms. A high percentage of the human hematopoietic cell chimerism persisted for more than 24 weeks after transplantation, and hCD34(+) bone marrow grafts of primary recipients could reconstitute hematopoiesis in secondary NOD/SCID/IL2rgamma(null) recipients, suggesting that this system can support self-renewal of human hematopoietic stem cells. hCD34(+)hCD38(-) CB cells differentiated into mature blood cells, including myelomonocytes, dendritic cells, erythrocytes, platelets, and lymphocytes. Differentiation into each lineage occurred via developmental intermediates such as common lymphoid progenitors and common myeloid progenitors, recapitulating the steady-state human hematopoiesis. B cells underwent normal class switching, and produced antigen-specific immunoglobulins (Igs). T cells displayed the human leukocyte antigen (HLA)-dependent cytotoxic function. Furthermore, human IgA-secreting B cells were found in the intestinal mucosa, suggesting reconstitution of human mucosal immunity. Thus, the NOD/SCID/IL2rgamma(null) newborn system might be an important experimental model to study the human hemato-lymphoid system.

ADP-ribosyl Cyclase↗

Distinctive and indispensable roles of PU.1 in maintenance of hematopoietic stem cells and their differentiation.

The PU.1 transcription factor is a key regulator of hematopoietic development, but its role at each hematopoietic stage remains unclear. In particular, the expression of PU.1 in hematopoietic stem cells (HSCs) could simply represent "priming" of genes related to downstream myelolymphoid lineages. By using a conditional PU.1 knock-out model, we here show that HSCs express PU.1, and its constitutive expression is necessary for maintenance of the HSC pool in the bone marrow. Bone marrow HSCs disrupted with PU.1 in situ could not maintain hematopoiesis and were outcompeted by normal HSCs. PU.1-deficient HSCs also failed to generate the earliest myeloid and lymphoid progenitors. PU.1 disruption in granulocyte/monocyte-committed progenitors blocked their maturation but not proliferation, resulting in myeloblast colony formation. PU.1 disruption in common lymphoid progenitors, however, did not prevent their B-cell maturation. In vivo disruption of PU.1 in mature B cells by the CD19-Cre locus did not affect B-cell maturation, and PU.1-deficient mature B cells displayed normal proliferation in response to mitogenic signals including the cross-linking of surface immunoglobulin M (IgM). Thus, PU.1 plays indispensable and distinct roles in hematopoietic development through supporting HSC self-renewal as well as commitment and maturation of myeloid and lymphoid lineages.

Animals↗

The complex cartography of stem cell commitment.

In this issue of Cell, a study by Adolfsson and coworkers (Adolfsson et al., 2005) provides insight into the early lineage commitment events of multipotent hematopoietic stem cells (HSCs). These studies demonstrate the importance of the Flt3 receptor tyrosine kinase as the earliest marker of hematopoietic cell fate commitment in that erythrocyte and megakaryocyte potentials are lost first as HSCs differentiate to lymphocyte progenitors.

Animals↗

Loss of Runx1 perturbs adult hematopoiesis and is associated with a myeloproliferative phenotype.

Homozygous loss of function of Runx1 (Runt-related transcription factor 1 gene) during murine development results in an embryonic lethal phenotype characterized by a complete lack of definitive hematopoiesis. In light of recent reports of disparate requirements for hematopoietic transcription factors during development as opposed to adult hematopoiesis, we used a conditional gene-targeting strategy to effect the loss of Runx1 function in adult mice. In contrast with the critical role of Runx1 during development, Runx1 was not essential for hematopoiesis in the adult hematopoietic compartment, though a number of significant hematopoietic abnormalities were observed. Runx1 excision had lineage-specific effects on B- and T-cell maturation and pronounced inhibition of common lymphocyte progenitor production. Runx1 excision also resulted in inefficient platelet production. Of note, Runx1-deficient mice developed a mild myeloproliferative phenotype characterized by an increase in peripheral blood neutrophils, an increase in myeloid progenitor populations, and extramedullary hematopoiesis composed of maturing myeloid and erythroid elements. These findings indicate that Runx1 deficiency has markedly different consequences during development compared with adult hematopoiesis, and they provide insight into the phenotypic manifestations of Runx1 deficiency in hematopoietic malignancies.

Animals↗

Obligate role of anti-apoptotic MCL-1 in the survival of hematopoietic stem cells.

Apoptosis is important in controlling hematopoietic stem cell (HSC) numbers. However, the specific BCL-2 family member(s) that regulate HSC homeostasis are not precisely defined. We tested myeloid leukemia-1 (MCL-1) as an attractive candidate that is highly expressed in HSCs and regulated by growth factor signals. Inducible deletion of Mcl-1 in mice resulted in ablation of bone marrow. This resulted in the loss of early bone marrow progenitor populations, including HSCs. Moreover, growth factors including stem cell factor increased transcription of the Mcl-1 gene and required MCL-1 to augment survival of purified bone marrow progenitors. Deletion of Mcl-1 in other tissues, including liver, did not impair survival. Thus, MCL-1 is a critical and specific regulator essential for ensuring the homeostasis of early hematopoietic progenitors.

Animals↗

TEF, an antiapoptotic bZIP transcription factor related to the oncogenic E2A-HLF chimera, inhibits cell growth by down-regulating expression of the common beta chain of cytokine receptors.

Gain and/or loss of function mediated by chimeric transcription factors generated by nonrandom translocations in leukemia is a key to understanding oncogenesis. E2A-hepatic leukemia factor (HLF), a chimeric basic region/leucine zipper (bZIP) transcription factor expressed in t(17;19)-positive leukemia cells, contributes to leukemogenesis through its potential to inhibit apoptosis. To identify physiologic counterparts of this chimera, we investigated the function of other bZIP factors that bind to the same DNA sequence recognized by E2A-HLF. Here, we show that thyrotroph embryonic factor (TEF), which shares a high level of sequence identity with HLF and recognizes the same DNA sequence, is expressed in a small fraction of each subset of hematolymphoid progenitors. When TEF was introduced into FL5.12 interleukin 3 (IL-3)-dependent cells, TEF protected the cells from apoptosis due to IL-3 deprivation. Unexpectedly, TEF also almost completely down-regulated expression of the common beta (betac) chain of cytokine receptors. Consequently, TEF-expressing cells accumulated in G(0)/G(1) phase without undergoing apoptosis. These findings suggest that TEF is one of the apoptotic regulators in hematopoietic progenitors and controls hematopoietic-cell proliferation by regulating the expression of the betac chain. In contrast, E2A-HLF promoted cell survival more efficiently than TEF but did not down-regulate betac chain expression, suggesting that E2A-HLF retains ideal properties for driving leukemic transformation.

Animals↗

Conditional MLL-CBP targets GMP and models therapy-related myeloproliferative disease.

Chromosomal translocations that fuse the mixed lineage leukemia (MLL) gene with multiple partners typify acute leukemias of infancy as well as therapy-related leukemias. We utilized a conditional knockin strategy to bypass the embryonic lethality caused by MLL-CBP expression and to assess the immediate effects of induced MLL-CBP expression on hematopoiesis. Within days of activating MLL-CBP, the fusion protein selectively expanded granulocyte/macrophage progenitors (GMP) and enhanced their self-renewal/proliferation. MLL-CBP altered the gene expression program of GMP, upregulating a subset of genes including Hox a9. Inhibition of Hox a9 expression by RNA interference demonstrated that MLL-CBP required Hox a9 for its enhanced cell expansion. Following exposure to sublethal gamma-irradiation or N-ethyl-N-nitrosourea (ENU), MLL-CBP mice developed myelomonocytic hyperplasia and progressed to fatal myeloproliferative disorders. These represented the spectrum of therapy-induced acute myelomonocytic leukemia/chronic myelomonocytic leukemia/myelodysplastic/myeloproliferative disorder similar to that seen in humans possessing the t(11;16). This model of MLL-CBP therapy-related myeloproliferative disease demonstrates the selectivity of this MLL fusion for GMP cells and its ability to initiate leukemogenesis in conjunction with cooperating mutations.

Animals↗

Prognostic, therapeutic, and mechanistic implications of a mouse model of leukemia evoked by Shp2 (PTPN11) mutations.

The SH2-containing tyrosine phosphatase Shp2 (PTPN11) is required for growth factor and cytokine signaling. Germline Shp2 mutations cause Noonan Syndrome (NS), which is associated with increased risk of juvenile myelomonocytic leukemia (JMML). Somatic Shp2 mutations occur in sporadic JMML and other leukemias. We found that Shp2 mutants associated with sporadic leukemias transform murine bone marrow cells, whereas NS mutants are less potent in this assay. Transformation requires multiple domains within Shp2 and the Shp2 binding protein Gab2, and is associated with hyperactivation of the Erk, Akt, and Stat5 pathways. Mutant Shp2-transduced BM causes a fatal JMML-like disorder or, less commonly, lymphoproliferation. Shp2 mutants also cause myeloproliferation in Drosophila. Mek or Tor inhibitors potently inhibit transformation, suggesting new approaches to JMML therapy.

Alleles↗

Potential autoregulation of transcription factor PU.1 by an upstream regulatory element.

Regulation of the hematopoietic transcription factor PU.1 (Spi-1) plays a critical role in the development of white cells, and abnormal expression of PU.1 can lead to leukemia. We previously reported that the PU.1 promoter cannot induce expression of a reporter gene in vivo, and cell-type-specific expression of PU.1 in stable lines was conferred by a 3.4-kb DNA fragment including a DNase I hypersensitive site located 14 kb upstream of the transcription start site. Here we demonstrate that this kb -14 site confers lineage-specific reporter gene expression in vivo. This kb -14 upstream regulatory element contains two 300-bp regions which are highly conserved in five mammalian species. In Friend virus-induced erythroleukemia, the spleen focus-forming virus integrates into the PU.1 locus between these two conserved regions. DNA binding experiments demonstrated that PU.1 itself and Elf-1 bind to a highly conserved site within the proximal homologous region in vivo. A mutation of this site abolishing binding of PU.1 and Elf-1 led to a marked decrease in the ability of this upstream element to direct activity of reporter gene in myelomonocytic cell lines. These data suggest that a potential positive autoregulatory loop mediated through an upstream regulatory element is essential for proper PU.1 gene expression.

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