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Cited by 63Genomics and Chromatin DynamicsAcute Myeloid Leukemia ResearchRetinoids in leukemia and cellular processes

RUNX1 regulates corepressor interactions of PU.1

Open access
Hu, Zhenbo, Gu, Xiaorong, Baraoidan, Kristine, Ibanez, Vinzon, Sharma, Arun, Kadkol, ShriHari, Munker, Reinhold, Ackerman, Steven +2 more·2011·Blood
DOI 10.1182/blood-2010-10-312512PMC PMC3123020
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Abstract

The transcription factor (TF) RUNX1 cooperates with lineage-specifying TFs (eg, PU.1/SPI1) to activate myeloid differentiation genes, such as macrophage and granulocyte macrophage colony-stimulating factor receptors (MCSFR and GMCSFR). Disruption of cooperative gene activation could contribute to aberrant repression of differentiation genes and leukemogenesis initiated by mutations and translocations of RUNX1. To investigate the mechanisms underlying cooperative gene activation, the effects of Runx1 deficiency were examined in an in vitro model of Pu.1-driven macrophage differentiation and in primary cells. Runx1 deficiency decreased Pu.1-mediated activation of Mcsfr and Gmcsfr, accompanied by decreased histone acetylation at the Mcsfr and Gmcsfr promoters, and increased endogenous corepressor (Eto2, Sin3A, and Hdac2) coimmunoprecipitation with Pu.1. In cotransfection experiments, corepressors were excluded from a multiprotein complex containing full-length RUNX1 and PU.1. However, corepressors interacted with PU.1 if wild-type RUNX1 was replaced with truncated variants associated with leukemia. Histone deacetylase (HDAC) enzyme activity is a major component of corepressor function. HDAC inhibition using suberoylanilide hydroxamic acid or MS-275 significantly increased MCSFR and GMCSFR expression in leukemia cell lines that express PU.1 and mutated or translocated RUNX1. RUNX1 deficiency is associated with persistent corepressor interaction with PU.1. Thus, inhibiting HDAC can partly compensate for the functional consequences of RUNX1 deficiency.

AI snapshot

Written from the paper’s full text.

TL;DR

RUNX1 and PU.1 cooperate to exclude corepressors from a transcription factor complex, enabling activation of myeloid differentiation genes such as MCSFR and GMCSFR. RUNX1 deficiency or leukemia-associated RUNX1 truncation permits persistent corepressor interaction with PU.1, and HDAC inhibition partly restores MCSFR/GMCSFR expression.

Key findings

  • Runx1 suppression in PUER cells (>50% by quantitative RT-PCR and Western blot) impaired Pu.1-mediated activation of Mcsfr, Gmcsfr, and F4/80 while preserving repression of stem cell genes HoxB4, Bmi-1, and c-Kit.
  • Runx1 deficiency increased coimmunoprecipitation of endogenous corepressors Eto2, Sin3a, and Hdac2 with Pu.1; LC-MS/MS detected Hdac2 unique peptides 2 vs 6 and Sin3a none detected vs 2 unique peptides in control vs Runx1-deficient PUER cells.
  • Full-length RUNX1 excluded ETO2 and SIN3A from the RUNX1/PU.1 complex, whereas leukemia-associated RUNX1-ETO or RUNX1ΔC-terminus did not.
  • HDAC inhibitors MS-275 (0.5 μM) and SAHA (1 μM) significantly increased MCSFR and GMCSFR mRNA and protein in Kasumi-1 and CG-SH AML cells and increased Mcsfr/Gmcsfr in primary murine Runx1+/− hematopoietic cells.

Methods

  • In vitro PUER murine hematopoietic precursor cells with shRNA-mediated Runx1 suppression and OHT-triggered Pu.1 macrophage differentiation; primary murine Runx1+/− and wild-type littermate bone marrow cells; human AML cell lines Kasumi-1 and CG-SH.
  • Quantitative RT-PCR, ChIP for histone H3 acetylation at Mcsfr/Gmcsfr/Gcsfr promoters, coimmunoprecipitation/Western blot, LC-MS/MS, transient transfection with RUNX1, RUNX1-ETO, RUNX1ΔC-terminus, PU.1, and ETO2 vectors.
  • Public microarray dataset GSE14471 analyzed for PU.1, CEBPA, MCSFR, and GMCSFR expression; HDAC inhibitor treatment with MS-275 and SAHA.

Limitations

  • Not stated in the paper: the paper does not include a dedicated limitations section.
  • Evident from the design: mechanistic conclusions rely mainly on PUER cell lines, murine Runx1+/− primary bone marrow, and human AML cell lines, with primary human AML evidence limited to analysis of a public microarray dataset.
  • Evident from the design: HDAC inhibitor rescue was tested in cell lines and murine Runx1+/− primary cells, not in primary human AML cells or in vivo leukemia models, and no clinical outcomes are reported.

Model-written summary. Check it against the paper before relying on it.

Integrity signals

Registry

Crossref · 10.1182/blood-2010-10-312512

Cites

50 of 50 references verified0 citation errors · 0 not checkable

Statistics

Nothing to recompute.

Claims the paper makes

30 of 90 cited sources checked · 15 supported · 0 flagged · accuracy not yet measured

  • Premiseown data
    § Abstract

    RUNX1 cooperates with lineage-specifying TFs (e.g., PU.1/SPI1) to activate myeloid differentiation genes such as MCSFR and GMCSFR.

    “The transcription factor (TF) RUNX1 cooperates with lineage-specifying TFs (eg, PU.1/SPI1) to activate myeloid differentiation genes, such as macrophage and granulocyte macrophage colony-stimulating factor receptors (MCSFR and GMCSFR).”
    Quote in the paperSubstantial claim3 of 7 sources checked
    • cites [4] Elagib, 2003, RUNX1 and GATA-1 coexpression and cooperation in megakaryocytic differentiation., Blood source
    • cites [5] Waltzer, 2003, Cooperation between the GATA and RUNX factors Serpent and Lozenge during Drosophila hematopoiesis., EMBO J source
    • cites [6] Huang, 2009, Differentiation-dependent interactions between RUNX-1 and FLI-1 during megakaryocyte development., Mol Cell Biol source

      The Discussion (page unavailable) says that RUNX-1 interactions with PU.1 have been previously described, supporting the cooperation aspect. The Results (page unavailable) instead examine RUNX-1 and FLI-1 regulation of megakaryocyte-specific genes; the paper does not establish that RUNX-1 and PU.1 activate myeloid differentiation genes MCSFR or GMCSFR. (LLM, conf=0.86)

      Experimental — accuracy not yet measured
    • cites [7] Zhang, 1996, CCAAT enhancer-binding protein (C/EBP) and AML1 (CBF alpha2) synergistically activate the macrophage colony-stimulating factor receptor promoter., Mol Cell Biol source
    • cites [8] Petrovick, 1998, Multiple functional domains of AML1: PU.1 and C/EBPalpha synergize with different regions of AML1., Mol Cell Biol source

      The section on PU.1–AML1 synergy reports that AML1B (RUNX1) and PU.1 together activate the M-CSF receptor promoter more than additively. The paper does not show their cooperation in activating the GM-CSF receptor gene; its mention of GM-CSF concerns the cytokine gene, not its receptor. The cited result also concerns promoter activity in transfected cells rather than direct measurement of differentiation. (LLM, conf=0.89)

      Experimental — accuracy not yet measured
    • cites [9] Kim, 1999, Mutual activation of Ets-1 and AML1 DNA binding by direct interaction of their autoinhibitory domains., EMBO J source

      In “Mechanism of mutual activation of Ets-1 and PEBP2αB1” (page unavailable), the paper says AML1/αB1 interacts and synergizes with PU.1 in activating the macrophage colony-stimulating factor receptor promoter. It does not say that AML1 and PU.1 cooperate to activate the granulocyte-macrophage colony-stimulating factor receptor; the References name a study of that receptor involving PU.1 and C/EBPα instead. (LLM, conf=0.88)

      Experimental — accuracy not yet measured
    • cites [10] Goetz, 2000, Autoinhibition of Ets-1 is counteracted by DNA binding cooperativity with core-binding factor alpha2., Mol Cell Biol source
  • Premisecitation
    § Introduction

    RUNX factors alone are relatively weak activators of transcription.

    “(RUNX factors alone are relatively weak activators of transcription.4,5,7,8,11)”
    Quote in the paperSubstantial claim2 of 5 sources checked
    • cites [4] Elagib, 2003, RUNX1 and GATA-1 coexpression and cooperation in megakaryocytic differentiation., Blood source
    • cites [5] Waltzer, 2003, Cooperation between the GATA and RUNX factors Serpent and Lozenge during Drosophila hematopoiesis., EMBO J source
    • cites [7] Zhang, 1996, CCAAT enhancer-binding protein (C/EBP) and AML1 (CBF alpha2) synergistically activate the macrophage colony-stimulating factor receptor promoter., Mol Cell Biol source
    • cites [8] Petrovick, 1998, Multiple functional domains of AML1: PU.1 and C/EBPalpha synergize with different regions of AML1., Mol Cell Biol source

      In the section on PU.1–AML1 synergy (page not available), AML1B alone activated the M-CSF receptor promoter 3.5-fold in HeLa cells, versus 12-fold with PU.1. This supports relatively modest activation by AML1B alone in that assay, but does not establish the claim for RUNX factors generally. (LLM, conf=0.83)

      Experimental — accuracy not yet measured
    • cites [11] Puig-Kroger, 2003, RUNX/AML and C/EBP factors regulate CD11a integrin expression in myeloid cells through overlapping regulatory elements., Blood source
  • Premisecitation
    § Introduction

    RUNX1 synergistically increases transcriptional activation by ETS1, PU.1(SPI1), CEBPA, GATA1, GATA2, and FLI1.

    “RUNX1 synergistically increases transcriptional activation by ETS1, PU.1(SPI1), CCAAT/enhancer binding protein-α (CEBPA), GATA1, GATA2, and FLI1.4–10”
    Quote in the paperSubstantial claim3 of 7 sources checked
    • cites [4] Elagib, 2003, RUNX1 and GATA-1 coexpression and cooperation in megakaryocytic differentiation., Blood source
    • cites [5] Waltzer, 2003, Cooperation between the GATA and RUNX factors Serpent and Lozenge during Drosophila hematopoiesis., EMBO J source
    • cites [6] Huang, 2009, Differentiation-dependent interactions between RUNX-1 and FLI-1 during megakaryocyte development., Mol Cell Biol source
    • cites [7] Zhang, 1996, CCAAT enhancer-binding protein (C/EBP) and AML1 (CBF alpha2) synergistically activate the macrophage colony-stimulating factor receptor promoter., Mol Cell Biol source
    • cites [8] Petrovick, 1998, Multiple functional domains of AML1: PU.1 and C/EBPalpha synergize with different regions of AML1., Mol Cell Biol source

      The Results report synergistic activation of the M-CSF receptor promoter by AML1 (RUNX1) with PU.1, and the paper also reports synergy with C/EBPα. The Discussion describes cooperative binding with ETS1, but does not establish the claimed transcriptional synergy with ETS1; our copy provides no support for the GATA1, GATA2, or FLI1 parts of the list. Page numbers were not available in the indexed passages. (LLM, conf=0.88)

      Experimental — accuracy not yet measured
    • cites [9] Kim, 1999, Mutual activation of Ets-1 and AML1 DNA binding by direct interaction of their autoinhibitory domains., EMBO J source

      The section on cooperative transactivation reports that AML1 (αB1) and Ets-1 synergistically activate the TCRβ enhancer. The mechanism section also says αB1 synergizes with PU.1 and C/EBPα in transactivation of a macrophage colony-stimulating factor receptor promoter. The passages provide no corresponding support for GATA1, GATA2, or FLI1. Page numbers are unavailable for these passages in our copy. (LLM, conf=0.88)

      Experimental — accuracy not yet measured
    • cites [10] Goetz, 2000, Autoinhibition of Ets-1 is counteracted by DNA binding cooperativity with core-binding factor alpha2., Mol Cell Biol source
  • Premisecitation
    § Introduction

    The specific molecular mechanisms underlying RUNX1 cooperation with lineage-specifying TFs have only been well characterized for RUNX1 cooperation with ETS1.

    “Although the aforementioned evidence suggests that cooperation between RUNX1 and lineage-specifying TF is an important aspect of normal and leukemic hematopoiesis, the specific molecular mechanisms underlying such cooperation have only been well characterized for RUNX1 cooperation with ETS1.9,10”
    Quote in the paperSubstantial claim1 of 2 sources checked
    • cites [9] Kim, 1999, Mutual activation of Ets-1 and AML1 DNA binding by direct interaction of their autoinhibitory domains., EMBO J source

      The Discussion and “Mechanism of mutual activation of Ets-1 and PEBP2αB1” characterize how AML1 (αB1) and Ets-1 interact through autoinhibitory domains to activate DNA binding. The paper discusses cooperation with other transcription factors, but does not establish the broader claim that ETS1 is the only RUNX1 partner for which a molecular mechanism has been well characterized. (LLM, conf=0.82)

      Experimental — accuracy not yet measured
    • cites [10] Goetz, 2000, Autoinhibition of Ets-1 is counteracted by DNA binding cooperativity with core-binding factor alpha2., Mol Cell Biol source
  • Premisecitation
    § Results

    In cotransfection experiments, RUNX1, PU.1, and CEBPA play essential roles in activation of the hematopoietic differentiation genes MCSFR and GMCSFR.

    “In cotransfection experiments, RUNX1, PU.1, and CEBPA play essential roles in activation of the hematopoietic differentiation genes MCSFR and GMCSFR.7,8,31”
    Quote in the paperSubstantial claim2 of 3 sources checked
    • cites [7] Zhang, 1996, CCAAT enhancer-binding protein (C/EBP) and AML1 (CBF alpha2) synergistically activate the macrophage colony-stimulating factor receptor promoter., Mol Cell Biol source
    • cites [8] Petrovick, 1998, Multiple functional domains of AML1: PU.1 and C/EBPalpha synergize with different regions of AML1., Mol Cell Biol source

      The paper’s Abstract and its section on PU.1–AML1 synergy report transient-transfection evidence that AML1 (RUNX1), PU.1, and C/EBPα contribute to activation of the M-CSF receptor promoter. The cited paper does not show the three factors activating the GM-CSF receptor gene; its References instead list a separate study of PU.1 and C/EBPα regulation of that gene. Page numbers are unavailable in our copy. (LLM, conf=0.88)

      Experimental — accuracy not yet measured
    • cites [31] Hohaus, 1995, PU.1 (Spi-1) and C/EBP alpha regulate expression of the granulocyte-macrophage colony-stimulating factor receptor alpha gene., Mol Cell Biol source

      The Results report that cotransfected PU.1 and C/EBPα each increased GM-CSF receptor α promoter activity, supporting that part of the sentence. This paper does not report cotransfection evidence for RUNX1 or activation of the M-CSF receptor gene; it mentions the latter as another PU.1-regulated promoter. (LLM, conf=0.88)

      Experimental — accuracy not yet measured
  • May combine two claims. 27
  • Classified as probably not a substantial claim. 14
  • The quote only partly states this claim. 1

  • May combine two claims.

    Premisecitation
    § Introduction

    RUNX1 physically interacts with PU.1, CEBPA, GATA1, GATA2, PAX5, FLI1, and ETS1.

    “RUNX1 physically interacts with PU.1, CEBPA, GATA1, GATA2, PAX5, FLI1, and ETS1.4–10,13”
    Quote in the paperSubstantial claim4 of 8 sources checked
    • cites [4] Elagib, 2003, RUNX1 and GATA-1 coexpression and cooperation in megakaryocytic differentiation., Blood source
    • cites [5] Waltzer, 2003, Cooperation between the GATA and RUNX factors Serpent and Lozenge during Drosophila hematopoiesis., EMBO J source
    • cites [6] Huang, 2009, Differentiation-dependent interactions between RUNX-1 and FLI-1 during megakaryocyte development., Mol Cell Biol source

      The Discussion reports direct RUNX-1 interactions with PU.1 and ETS-1, as well as interactions with GATA-1 and GATA-2. The Results document RUNX-1–FLI-1 binding. The passages in our copy do not establish physical interactions with CEBPA or PAX5; searches for both found no supporting passage. Page numbers were not available in our copy. (LLM, conf=0.86)

      Experimental — accuracy not yet measured
    • cites [7] Zhang, 1996, CCAAT enhancer-binding protein (C/EBP) and AML1 (CBF alpha2) synergistically activate the macrophage colony-stimulating factor receptor promoter., Mol Cell Biol source
    • cites [8] Petrovick, 1998, Multiple functional domains of AML1: PU.1 and C/EBPalpha synergize with different regions of AML1., Mol Cell Biol source

      The Results and Discussion (page unavailable in our copy) state that AML1/RUNX1 physically interacts with PU.1 and C/EBPα. The Discussion mentions cooperative DNA binding with Ets-1, but does not establish a physical interaction with it. The passages do not support the claimed physical interactions with GATA1, GATA2, PAX5, or FLI1. (LLM, conf=0.90)

      Experimental — accuracy not yet measured
    • cites [9] Kim, 1999, Mutual activation of Ets-1 and AML1 DNA binding by direct interaction of their autoinhibitory domains., EMBO J source
    • cites [10] Goetz, 2000, Autoinhibition of Ets-1 is counteracted by DNA binding cooperativity with core-binding factor alpha2., Mol Cell Biol source
    • cites [13] Tagoh, 2006, The mechanism of repression of the myeloid-specific c-fms gene by Pax5 during B lineage restriction., EMBO J source

      The Results section mentions a Runx1/Egr-2/ets DNA-binding cluster, while the Figure 7 caption reports a complex containing Pax5 and PU.1. Neither establishes a physical interaction between RUNX1 and any of the listed proteins. Searches for RUNX1–Pax5 and other RUNX1 protein interactions found no passage supporting the attributed claim. (LLM, conf=0.76)

      Experimental — accuracy not yet measured

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation
    § Introduction

    The heterodimer β-subunit of RUNX1 and CBFB can relieve autoinhibition of RUNX1 DNA binding.

    “However, the heterodimer β-subunit of RUNX1 and CBFB can also relieve autoinhibition of RUNX1 DNA binding,16”
    Quote in the paperNot a substantial claim1 of 1 sources checked
    • cites [16] Kanno, 1998, Intrinsic transcriptional activation-inhibition domains of the polyomavirus enhancer binding protein 2/core binding factor alpha subunit revealed in the presence of the beta subunit., Mol Cell Biol source

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation
    § Introduction

    Both RUNX1 and ETS1 autoinhibit their own DNA binding; mutual interaction of their autoinhibitory domains relieves autoinhibition and leads to synergistic gene activation.

    “Both RUNX1 and ETS1 autoinhibit their own DNA binding.9,10 Mutual interaction of the autoinhibitory domains of RUNX1 and ETS1 mutually relieves the autoinhibition of DNA binding and leads to synergistic gene activation.9,10”
    Quote in the paperNot a substantial claim1 of 2 sources checked
    • cites [9] Kim, 1999, Mutual activation of Ets-1 and AML1 DNA binding by direct interaction of their autoinhibitory domains., EMBO J source
    • cites [10] Goetz, 2000, Autoinhibition of Ets-1 is counteracted by DNA binding cooperativity with core-binding factor alpha2., Mol Cell Biol source

  • May combine two claims.

    Interpretationreasoning
    § Discussion

    Loss of this function of the RUNX1 C-terminus could contribute to the inability of RUNT to synergize with PU.1 to transactivate the MCSFR promoter and to the impaired differentiation and leukemogenesis associated with RUNX1a expression.

    “Loss of this function of the RUNX1 C-terminus could contribute to the inability of RUNT to synergize with PU.1 to transactivate the MCSFR promoter,8 and the impaired differentiation and leukemogenesis associated with expression of RUNX1a (a RUNX1 isoform that lacks the RUNX1 C-terminus).38,39”
    Quote in the paperSubstantial claim2 of 3 sources checked
    • cites [8] Petrovick, 1998, Multiple functional domains of AML1: PU.1 and C/EBPalpha synergize with different regions of AML1., Mol Cell Biol source
    • cites [38] Tanaka, 1995, An acute myeloid leukemia gene, AML1, regulates hemopoietic myeloid cell differentiation and transcriptional activation antagonistically by two alternative spliced forms., EMBO J source
    • cites [39] Gutierrez-Angulo, 2005, Increased expression of AML1-a and acquired chromosomal abnormalities in childhood acute lymphoblastic leukemia., Hematol Oncol source

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation
    § Discussion

    RUNX1 alone is a relatively weak activator of transcription.

    “In keeping with this paradigm, RUNX1 alone is a relatively weak activator of transcription.4,5,7,8,11”
    Quote in the paperNot a substantial claim2 of 5 sources checked
    • cites [4] Elagib, 2003, RUNX1 and GATA-1 coexpression and cooperation in megakaryocytic differentiation., Blood source
    • cites [5] Waltzer, 2003, Cooperation between the GATA and RUNX factors Serpent and Lozenge during Drosophila hematopoiesis., EMBO J source
    • cites [7] Zhang, 1996, CCAAT enhancer-binding protein (C/EBP) and AML1 (CBF alpha2) synergistically activate the macrophage colony-stimulating factor receptor promoter., Mol Cell Biol source
    • cites [8] Petrovick, 1998, Multiple functional domains of AML1: PU.1 and C/EBPalpha synergize with different regions of AML1., Mol Cell Biol source
    • cites [11] Puig-Kroger, 2003, RUNX/AML and C/EBP factors regulate CD11a integrin expression in myeloid cells through overlapping regulatory elements., Blood source

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation
    § Discussion

    Runx1 deficiency permissiveness for partial differentiation has been seen in other models.

    “Runx1 deficiency permissiveness for partial differentiation has been seen in other models.”
    Quote in the paperNot a substantial claim1 of 1 sources checked
    • cites [28] Sun, 2004, Haploinsufficiency of AML1 results in a decrease in the number of LTR-HSCs while simultaneously inducing an increase in more mature progenitors., Blood source

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation
    § Discussion

    Hematopoietic precursors subject to a cytokine differentiation stimulus rapidly repress key stem cell genes.

    “Hematopoietic precursors subject to a cytokine differentiation stimulus rapidly repress key stem cell genes.44”
    Quote in the paperNot a substantial claim1 of 1 sources checked
    • cites [44] Hu, 2010, Decitabine maintains hematopoietic precursor self-renewal by preventing repression of stem cell genes by a differentiation-inducing stimulus., Mol Cancer Ther source

  • May combine two claims.

    Premisecitation
    § Discussion

    In Runx1+/− mice, hematopoietic stem cells are decreased but myeloid progenitors are increased.

    “In Runx1+/− mice, hematopoietic stem cells are decreased but myeloid progenitors are increased.28”
    Quote in the paperNot a substantial claim1 of 1 sources checked
    • cites [28] Sun, 2004, Haploinsufficiency of AML1 results in a decrease in the number of LTR-HSCs while simultaneously inducing an increase in more mature progenitors., Blood source

      The Discussion reports fewer long-term repopulating HSCs and more myeloid progenitors in AML1+/− mice. The Results specify that short-term repopulating HSCs did not differ significantly, so the claim's unqualified statement about hematopoietic stem cells is broader than the finding. (LLM, conf=0.87)

      Experimental — accuracy not yet measured

  • May combine two claims.

    Premisecitation
    § Discussion

    Treatment of normal hematopoietic stem cells with HDAC inhibitors maintains self-renewal, possibly by preventing repression of stem cell genes by differentiation stimuli.

    “In contrast, treatment of normal hematopoietic stem cells with HDAC inhibitors has the opposite effect, maintaining self-renewal,44,48–50 possibly by preventing repression of stem cell genes by differentiation stimuli.44”
    Quote in the paperNot a substantial claim2 of 4 sources checked
    • cites [44] Hu, 2010, Decitabine maintains hematopoietic precursor self-renewal by preventing repression of stem cell genes by a differentiation-inducing stimulus., Mol Cancer Ther source
      Experimental — accuracy not yet measured
    • cites [48] Milhem, 2004, Modification of hematopoietic stem cell fate by 5aza 2′deoxycytidine and trichostatin A., Blood source
    • cites [49] Bug, 2005, Valproic acid stimulates proliferation and self-renewal of hematopoietic stem cells., Cancer Res source

      The Abstract and Discussion (page not provided) report that valproic acid, an HDAC inhibitor, increases self-renewal and inhibits differentiation in normal hematopoietic stem cells. The Discussion proposes transcriptional reprogramming and altered signaling as possible mechanisms, but does not specifically say that treatment prevents differentiation stimuli from repressing stem-cell genes. (LLM, conf=0.84)

      Experimental — accuracy not yet measured
    • cites [50] De Felice, 2005, Histone deacetylase inhibitor valproic acid enhances the cytokine-induced expansion of human hematopoietic stem cells., Cancer Res source

  • May combine two claims.

    Findingown data
    § Results

    After addition of OHT to PUER control, HoxB4, Bmi-1, and c-Kit were repressed followed by activation of the macrophage differentiation genes Mcsfr, Gmcsfr, and F4/80.

    “After addition of OHT to PUER control, HoxB4, Bmi-1, and c-Kit were repressed followed by activation of the macrophage differentiation genes Mcsfr, Gmcsfr, and F4/80 (Figure 1A).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data
    § Results

    As PU.1 concentrations are increased, PU.1 preferentially coimmunoprecipitates with RUNX1 with a corresponding decrease in coimmunoprecipitated SIN3A.

    “However, as PU.1 concentrations are increased, PU.1 preferentially coimmunoprecipitates with RUNX1 with a corresponding decrease in coimmunoprecipitated SIN3A (Figure 3B, lanes 6, 9, and 12).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data
    § Results

    As PU.1 concentrations are increased by transient transfection, PU.1 preferentially coimmunoprecipitates with RUNX1 with a corresponding decrease in coimmunoprecipitated ETO2.

    “However, as PU.1 concentrations are increased in the cells by transient transfection, PU.1 preferentially coimmunoprecipitates with RUNX1 with a corresponding decrease in coimmunoprecipitated ETO2 (Figure 3A, lanes 6, 9, and 12).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data
    § Results

    In PUER control, OHT induced morphologic changes of macrophage differentiation and a decrease in proliferation.

    “In PUER control, OHT induced morphologic changes of macrophage differentiation (increase in cell size, decrease in nuclear:cytoplasmic ratio, clumping of nuclear chromatin, adherence to culture plates, cytoplasmic vacuolization), and a decrease in proliferation (Figure 1C-D).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data
    § Results

    Hdac2 peptides were detected in both immunoprecipitates but were substantially enriched in the immunoprecipitate from shRunx1 cells.

    “Although Hdac2 peptides were detected in both immunoprecipitates, these peptides were substantially enriched in the immunoprecipitate from shRunx1 cells (Table 1; supplemental Figure 4A).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data
    § Results

    In PUER shRunx1 treated with OHT, morphologic changes of macrophage differentiation did not occur, and the cells continued to expand exponentially.

    “However, in PUER shRunx1 treated with OHT, morphologic changes of macrophage differentiation did not occur (Figure 1D), and the cells continued to expand exponentially (Figure 1C), with”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data
    § Results

    Kasumi-1 and CG-SH cells express PU.1 and CEBPA.

    “Western blot was used to confirm that Kasumi-1 and CG-SH express PU.1 and CEBPA (Figure 5A).”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Findingown data
    § Results

    The Sin3a and Hdac2 bands were relatively weak compared with the Eto2 bands in the coimmunoprecipitation Western blots.

    “The Sin3a and Hdac2 bands were relatively weak compared with the Eto2 bands.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingreasoning
    § Discussion

    Runx1 deficiency allowed Pu.1-triggered repression events and instead selectively impaired activation of the differentiation genes Mcsfr, Gmcsfr, and F4/80.

    “Runx1 deficiency allowed these Pu.1-triggered repression events and instead selectively impaired activation of the differentiation genes Mcsfr, Gmcsfr, and F4/80.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Premisecitation
    § Introduction

    RUNX1 is necessary for definitive hematopoiesis; Runx1 knockout murine embryos have no detectable definitive erythrocytes or myeloid cells in circulation or livers and die in utero at E12.5.

    “RUNX1 is necessary for definitive hematopoiesis: Runx1 knockout murine embryos have no detectable definitive erythrocytes or myeloid cells in their circulation or livers and die in utero at embryonic day 12.5 (E12.5).3”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [3] Okuda, 1996, AML1, the target of multiple chromosomal translocations in human leukemia, is essential for normal fetal liver hematopoiesis., Cell source

  • May combine two claims.

    Interpretationown data
    § Introduction

    This cooperation is disrupted by RUNX1 deficiency or by RUNX1 variants associated with leukemia.

    “This cooperation is disrupted by RUNX1 deficiency or by RUNX1 variants associated with leukemia.”
    Quote in the paperSubstantial claim

  • Classified as probably not a substantial claim.

    Interpretationreasoning
    § Introduction

    In AML cells expressing mutated or translocated RUNX1 and wild-type PU.1, a possible alternative route to decreased activation of PU.1 target genes is disrupted RUNX1/PU.1 cooperation.

    “Therefore, in AML cells, which express mutated or translocated RUNX1 and wild-type PU.1, a possible alternative route to decreased activation of PU.1 target genes is disrupted RUNX1/PU.1 cooperation.”
    Quote in the paperSubstantial claim

  • Classified as probably not a substantial claim. May combine two claims.

    Premisecitation
    § Introduction

    PU.1 can interact with coactivators or corepressors to function as either a transcriptional activator or a transcriptional repressor.

    “PU.1 can interact with coactivators or corepressors to function as either a transcriptional activator or a transcriptional repressor.17–19”
    Quote in the paperSubstantial claim0 of 3 sources checked
    • cites [17] Suzuki, 2003, Direct association between PU.1 and MeCP2 that recruits mSin3A-HDAC complex for PU.1-mediated transcriptional repression., Oncogene source
    • cites [18] Yamamoto, 1999, Physical and functional interactions between the transcription factor PU.1 and the coactivator CBP., Oncogene source
    • cites [19] Kihara-Negishi, 2001, In vivo complex formation of PU.1 with HDAC1 associated with PU.1-mediated transcriptional repression., Oncogene source

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation
    § Results

    High PU.1 protein expression in AML cells, including primary M0 and M2 AML cells, has been previously reported.

    “High PU.1 protein expression in AML cells, including primary M0 and M2 AML cells, has been previously reported.15”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [15] Iida, 2000, Protein expression and constitutive phosphorylation of hematopoietic transcription factors PU.1 and C/EBP beta in acute myeloid leukemia blasts., Int J Hematol

  • Probably not a substantial claim: may not pass S2 (checkable), S3 (load-bearing).

    Interpretationreasoning
    § Introduction

    Other mechanisms for RUNX1 cooperation with lineage-specifying TFs probably operate.

    “Therefore, other mechanisms for cooperation probably operate.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Premisecitation
    § Introduction

    PU.1 is a key lineage-specifying TF; there is complete absence of macrophages, B cells, mature T cells, and a decrease in neutrophils and hematopoietic stem cells in Pu.1−/− mice.

    “PU.1 is a key lineage-specifying TF. There is a complete absence of macrophages, B cells, mature T cells, and a decrease in neutrophils and hematopoietic stem cells in Pu.1−/− mice.21”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [21] Scott, 1994, Requirement of transcription factor PU.1 in the development of multiple hematopoietic lineages., Science source

  • May combine two claims.

    Study design and conductown data
    § Results

    Runx1 suppression in PUER shRunx1 was more than 50% by quantitative RT-PCR and Western blot.

    “Runx1 suppression was more than 50% by quantitative RT-PCR (supplemental Figure 2A-B) and by Western blot (supplemental Figure 2C).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Premisecitation
    § Introduction

    In human AML, PU.1 mutations are not frequent, although they have been reported.

    “In human AML, however, PU.1 mutations are not frequent, although they have been reported.23–25”
    Quote in the paperSubstantial claim0 of 3 sources checked
    • cites [23] Lamandin, 2002, Are PU.1 mutations frequent genetic events in acute myeloid leukemia (AML)?, Blood source
    • cites [24] Vegesna, 2002, C/EBP-beta, C/EBP-delta, PU.1, AML1 genes: mutational analysis in 381 samples of hematopoietic and solid malignancies., Leuk Res source
    • cites [25] Mueller, 2002, Heterozygous PU.1 mutations are associated with acute myeloid leukemia., Blood source

  • May combine two claims.

    Premisecitation
    § Introduction

    PU.1 and CEBPA are expressed at high levels in leukemia cells containing mutated and translocated RUNX1, as shown by Western blot and publicly available microarray gene expression data.

    “Nonetheless, as shown by Western blot15 and in publicly available microarray gene expression data, PU.1 and CEBPA are expressed at high levels in leukemia cells containing mutated and translocated RUNX1.”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [15] Iida, 2000, Protein expression and constitutive phosphorylation of hematopoietic transcription factors PU.1 and C/EBP beta in acute myeloid leukemia blasts., Int J Hematol

  • The quote only partly states this claim.

    Interpretationown data
    § Discussion

    The present study describes cooperative exclusion of corepressors by two transcription factors, RUNX1 and PU.1.

    “The present study (summarized in Figure 6) describes cooperative exclusion of corepressors by 2 TFs.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationown data
    § Discussion

    The present observations suggest that truncated forms of RUNX1-ETO may continue to cause aberrant repression of late differentiation genes because loss of the RUNX1 C-terminus is sufficient to cause persistent corepressor recruitment to PU.1.

    “The present observations suggest that these truncated forms of RUNX1-ETO may continue to cause aberrant repression of late differentiation genes because loss of the RUNX1 C-terminus is sufficient to cause persistent corepressor recruitment to PU.1.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationown data
    § Discussion

    Corepressor exclusion required the RUNX1 C-terminus because RUNT and RUNX1-ETO, which lack the RUNX1 C-terminus, allowed corepressor interaction with PU.1.

    “Corepressor exclusion required the RUNX1 C-terminus because RUNT and the leukemia fusion protein RUNX1-ETO, which lacks the RUNX1 C-terminus, allowed corepressor interaction with PU.1.”
    Quote in the paperSubstantial claim

  • Classified as probably not a substantial claim.

    Premisecitation
    § Discussion

    The mechanisms that regulate corepressor versus coactivator interactions of RUNX1 and PU.1 are incompletely understood.

    “However, the mechanisms that regulate corepressor versus coactivator interactions of RUNX1 and PU.1 are incompletely understood.17–20”
    Quote in the paperSubstantial claim0 of 4 sources checked
    • cites [17] Suzuki, 2003, Direct association between PU.1 and MeCP2 that recruits mSin3A-HDAC complex for PU.1-mediated transcriptional repression., Oncogene source
    • cites [18] Yamamoto, 1999, Physical and functional interactions between the transcription factor PU.1 and the coactivator CBP., Oncogene source
    • cites [19] Kihara-Negishi, 2001, In vivo complex formation of PU.1 with HDAC1 associated with PU.1-mediated transcriptional repression., Oncogene source
    • cites [20] Chakraborty, 2003, SUV39H1 interacts with AML1 and abrogates AML1 transactivity: AML1 is methylated in vivo., Oncogene source

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation
    § Discussion

    The RUNX1 C-terminus is highly conserved from amino acids 291 to 411.

    “The RUNX1 C-terminus is highly conserved from amino acids 291 to 411.37”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [37] Coffman, 2003, Runx transcription factors and the developmental balance between cell proliferation and differentiation., Cell Biol Int source

  • Classified as probably not a substantial claim.

    Premisecitation
    § Results

    The chromosomal translocation t(8;21) is a recurrent abnormality in AML that replaces the RUNX1 C-terminal transactivation domain with most of ETO.

    “The chromosomal translocation t(8;21) is a recurrent abnormality in AML that replaces the RUNX1 C-terminal transactivation domain with most of ETO.2”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [2] Miyoshi, 1991, t(8;21) breakpoints on chromosome 21 in acute myeloid leukemia are clustered within a limited region of a single gene, AML1., Proc Natl Acad Sci U S A source

  • May combine two claims.

    Interpretationreasoning
    § Discussion

    These results suggest a molecular mechanism for cooperative differentiation gene activation by RUNX1/PU.1 and for aberrant repression of late differentiation genes in AML cells containing mutated or translocated RUNX1.

    “These results suggest a molecular mechanism for cooperative differentiation gene activation by RUNX1/PU.1, and for aberrant repression of late differentiation genes in AML cells containing mutated or translocated RUNX1.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationreasoning
    § Discussion

    These observations provide a mechanistic explanation for the effect of HDAC inhibitors and augment the rationale for development of corepressor inhibitors as differentiation therapy agents for myelodysplastic syndrome and AML.

    “These observations provide a mechanistic explanation for this effect of HDAC inhibitors and augment the rationale for development of corepressor inhibitors as differentiation therapy agents for myelodysplastic syndrome and AML.”
    Quote in the paperNot a substantial claim

  • Classified as probably not a substantial claim. May combine two claims.

    Premisecitation
    § Discussion

    Physiologic gene activation is most probably cooperative involving multiple transcription factors collaborating to bind DNA, exclude corepressors, recruit coactivators, and recruit the basal transcription factor complex.

    “Physiologic gene activation is most probably cooperative involving multiple TFs collaborating to bind DNA, exclude corepressors, recruit coactivators, and recruit the basal TF complex.32”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [32] Carey, 1998, The enhanceosome and transcriptional synergy., Cell source

  • May combine two claims.

    Premisecitation
    § Discussion

    Transcripts from these mutated RUNX1 genes can escape nonsense-mediated decay, resulting in expression of RUNX1 variants that lack the C-terminus.

    “Transcripts from these mutated RUNX1 genes can escape nonsense-mediated decay, resulting in expression of RUNX1 variants that lack the C-terminus.43”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [43] Heller, 2005, Low Mpl receptor expression in a pedigree with familial platelet disorder with predisposition to acute myelogenous leukemia and a novel AML1 mutation., Blood source

  • May combine two claims.

    Premisecitation
    § Discussion

    In primary AML cells with translocated RUNX1, there is high expression of PU.1 and CEBPA but relatively low expression of MCSFR and GMCSFR.

    “In primary AML cells with translocated RUNX1, there is high expression of PU.115 and CEBPA (indicating lineage-commitment of the cells), but relatively low expression of MCSFR and GMCSFR.”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [15] Iida, 2000, Protein expression and constitutive phosphorylation of hematopoietic transcription factors PU.1 and C/EBP beta in acute myeloid leukemia blasts., Int J Hematol
39 of 39

  • The paper relies on this without support.

    RUNX1 is insufficient for hematopoiesis; hematopoietic lineage specification and differentiation require and are driven by key lineage-specifying transcription factors such as ETS, CEBP, and GATA families.

    “However, RUNX1 is insufficient for hematopoiesis; hematopoietic lineage specification and differentiation require and are driven by key lineage-specifying transcription factors (TFs), such as members of the ETS (including PU.1), CEBP, and GATA families.”

  • The paper relies on this without support.

    A key determinant of gene activation versus repression by DNA-binding factors is coactivator versus corepressor recruitment.

    “A key determinant of gene activation versus repression by DNA-binding factors is coactivator versus corepressor recruitment.”

  • The paper relies on this without support.

    Histone deacetylase (HDAC) enzyme activity is a major aspect of corepressor function.

    “Histone deacetylase (HDAC) enzyme activity is a major aspect of corepressor function.”

  • The paper relies on this without support.

    Corepressors Eto2 and Sin3a have a well-established role in hematopoiesis and associate with Hdac2; Hdac2 is highly expressed in hematopoietic cells.

    “Corepressors with a well-established role in hematopoiesis include Eto2 and Sin3a, both of which associate with Hdac2 (Hdac2 is an HDAC that is highly expressed in hematopoietic cells).”

  • The paper relies on this without support.

    RUNX1 and PU.1 physically interact with each other and with corepressors.

    “RUNX1 and PU.1 physically interact with each other and with corepressors.”

  • The paper relies on this without support.

    HDAC enzymatic activity is a major component of corepressor function.

    “HDAC enzymatic activity is a major component of corepressor function.”
6 of 6

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Authors
Hu, Z., Gu, X., Baraoidan, K., Ibanez, V., Sharma, A., Kadkol, S., Munker, R., Ackerman, S., Nucifora, G., & Saunthararajah, Y.
Affiliations
  • Cleveland Clinic (US)
  • University of Illinois Chicago (US)
  • Department of Medicine, University of Illinois at Chicago, Chicago, IL;
  • Northwestern University (US)
  • Louisiana State University Health Sciences Center Shreveport (US)
ORCID
Venue
Blood
Publisher
American Society of Hematology
Published
2011-06-16
Volume
117
Issue
24
Pages
6498-6508
ISSN
  • 0006-4971 (ISSN-L)
  • 1528-0020
DOI
10.1182/blood-2010-10-312512
PMC
PMC3123020
OpenAlex
W2067970911
Cited by
63
References
52
Citations by year
2026: 5 · 2024: 1 · 2023: 4 · 2022: 6 · 2021: 9 · 2020: 6 · 2019: 1 · 2018: 5 · 2017: 5 · 2016: 2 · 2015: 6 · 2014: 1 · 2013: 3 · 2012: 4
FWCI
1.07
Type
journal article
Language
en
Open access
Locations
  • Blood (publishedVersion)
  • PubMed (publishedVersion)
  • Europe PMC (PubMed Central) (submittedVersion)
  • PubMed Central (submittedVersion)
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Genomics and Chromatin Dynamics
  • Health Sciences › Medicine › Hematology › Acute Myeloid Leukemia Research
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Retinoids in leukemia and cellular processes
Keywords
RUNX1, PU.1, histone deacetylase, histone acetylation, macrophage colony-stimulating factor receptor, myeloid differentiation, leukemogenesis, vorinostat
MeSH terms
Proto-Oncogene Protein Spi-1, Animals, Humans, Mice, Transgenic, Protein Binding, Proto-Oncogene Proteins, Transfection, Trans-Activators

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Registry, retraction, Cites and statistics results are computed, not judged, with one exception: a printed reference that no index lists is read once by a model before it counts as a citation error. Claims are extracted by a model and kept only when their quote appears verbatim in the text; each cited source is then checked, with a model’s help, for whether it says what the claim relies on. Those verdicts can be wrong — read the claim’s quote and the cited source before relying on one.

Claims read by deepseek-v4.1-flash under definition version 3 (extract-v3) on 2026-10-07.

Classified by jev-1.13-20260917 (gold-guideline-v0.2) on 2026-10-07.

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