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Cited by 71Genomics and Chromatin DynamicsAcute Myeloid Leukemia ResearchEpigenetics and DNA Methylation

Inhibition of CBP-Mediated Protein Acetylation by the Ets Family Oncoprotein PU.1

Open access
Hong, Wei, Kim, Alexander Y., Ky, Sokun, Rakowski, Carrie, Seo, Sang-Beom, Chakravarti, Debabrata, Atchison, Michael, Blobel, Gerd A.·2002·Molecular and Cellular Biology
DOI 10.1128/mcb.22.11.3729-3743.2002PMC PMC133832
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Abstract

Aberrant expression of PU.1 inhibits erythroid cell differentiation and contributes to the formation of murine erythroleukemias (MEL). The molecular mechanism by which this occurs is poorly understood. Here we show that PU.1 specifically and efficiently inhibits CBP-mediated acetylation of several nuclear proteins, including the hematopoietic transcription factors GATA-1, NF-E2, and erythroid Krüppel-like factor. In addition, PU.1 blocks acetylation of histones and interferes with acetylation-dependent transcriptional events. CBP acetyltransferase activity increases during MEL cell differentiation as PU.1 levels decline and is inhibited by sustained PU.1 expression. Finally, PU.1 inhibits the differentiation-associated increase in histone acetylation at an erythroid-specific gene locus in vivo. Together, these findings suggest that aberrant expression of PU.1 and possibly other members of the Ets family of oncoproteins subverts normal cellular differentiation in part by inhibiting the acetylation of critical nuclear factors involved in balancing cellular proliferation and maturation.

AI snapshot

Written from the paper’s full text.

TL;DR

PU.1 inhibits CBP-mediated acetylation of GATA-1, other hematopoietic transcription factors, p53, and histones in vitro and in cells, and blocks acetylation-dependent transcription. Sustained PU.1 expression prevents differentiation-associated increases in CBP acetyltransferase activity and histone acetylation at the β-globin locus, suggesting a mechanism for PU.1-driven erythroid differentiation arrest.

Key findings

  • PU.1 dose-dependently inhibited CBP-mediated GATA-1 acetylation in vitro, with an IC50 of approximately 185 nM for full-length GATA-1 and almost complete inhibition at fivefold molar excess over GATA-1. It also inhibited acetylation of EKLF (IC50 ~600 nM), MafG (IC50 ~125 nM), p53, and histones (IC50 ~110 nM), but did not inhibit CBP autoacetylation.
  • PU.1 inhibited GATA-1 acetylation in transfected 3T3 cells and reduced acetylation of endogenous proteins in 293 cells. It also inhibited transcriptional activity of NF-E2, EKLF, and p53, and blocked AT-dependent GAL4-CBP-AT activation.
  • In MEL cells, GAL4-CBP-AT activity increased from 4–5-fold in undifferentiated cells to up to 17-fold after DMSO-induced differentiation; sustained PU.1 expression blocked this increase and blocked differentiation as measured by β-globin expression and benzidine staining.
  • ChIP assays showed differentiation-induced histone H3 acetylation increased approximately twofold at the β-major globin promoter and HS3 in control MEL cells but was completely inhibited in MEL-PU.1 cells; GAPDH acetylation did not increase. GATA-1 DNA binding was unchanged in MEL-PU.1 cells.

Methods

  • In vitro acetyltransferase assays used GST fusion proteins, purified CBP-AT, full-length baculovirus-expressed p300, PCAF, histones, and transcription factor substrates; acetylation was quantitated by phosphorimager analysis or scintillation counting.
  • Transient transfections were performed in NIH 3T3, 293, U2OS, Saos-2, and MEL cells, followed by immunoprecipitation/Western blotting with anti-acetyllysine and anti-GATA-1 antibodies, and luciferase reporter assays for NF-E2, EKLF, p53, and GAL4-CBP-AT.
  • Stable PU.1-expressing MEL cell lines were generated, induced with DMSO, and analyzed by RT-PCR for β-globin, benzidine staining, ChIP at the β-globin promoter/HS3/GAPDH, and gel shift assays for GATA-1 DNA binding.

Limitations

  • Paper-stated: the authors note it remains possible that PU.1 effects on CBP-AT activity in MEL cells are an indirect consequence of the differentiation block, although direct inhibition in transfected cells and GAL4-CBP-AT assays argue otherwise; the role of PU.1 interaction with RB in transformation remains to be seen; and the CBP sites to which PU.1 and EKLF bind have not been accurately mapped.
  • Evident from the text: many key assays rely on overexpression, transient transfection, or GST fusion proteins, and some structure-function data are described as preliminary. The study does not present an animal model or primary erythroleukemia transformation assay, and ChIP analyses are limited to selected loci.

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

Integrity signals

Registry

Crossref · 10.1128/mcb.22.11.3729-3743.2002

Cites

91 of 91 references verified0 citation errors · 0 not checkable

Statistics

Nothing to recompute.

Claims the paper makes

32 of 65 cited sources checked · 18 supported · 1 flagged · accuracy not yet measured

  • Premisecitation
    § Introduction

    Mutations in the acetylation sites impair the ability of GATA-1 to trigger differentiation of erythroid cells.

    “Mutations in the acetylation sites impair the ability of GATA-1 to trigger differentiation of erythroid cells (27), suggesting that acetylation is important for GATA-1 function in vivo.”
    Quote in the paperSubstantial claim1 of 1 sources checked
  • Premisecitation
    § RESULTS

    GAL4-CBP-AT activates a reporter gene containing GAL4-binding sites in transfected U2OS cells.

    “GAL4-CBP-AT activates a reporter gene containing GAL4-binding sites in transfected U2OS cells (39).”
    Quote in the paperSubstantial claim1 of 1 sources checked
  • Premisecitation
    § DISCUSSION

    These domains are also required for inhibition of MEL cell differentiation.

    “These domains are also required for inhibition of MEL cell differentiation (63).”
    Quote in the paperSubstantial claim1 of 1 sources checked
  • Premisecitation
    § RESULTS

    In the case of GATA-1, the acetylation status correlates with its transcriptional activity.

    “In the case of GATA-1, the acetylation status correlates with its transcriptional activity (27).”
    Quote in the paperSubstantial claim1 of 1 sources checked
  • Premisecitation
    § RESULTS

    Activation is dependent on AT activity, since a series of mutations across the AT domain revealed a tight correlation between AT activity and transcriptional output.

    “Activation is dependent on AT activity, since a series of mutations across the AT domain revealed a tight correlation between AT activity and transcriptional output (39).”
    Quote in the paperSubstantial claim1 of 1 sources checked
  • Classified as probably not a substantial claim. 32
  • May combine two claims. 35

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

    Premisecitation
    § DISCUSSION

    E1A can inhibit or stimulate CBP-AT activity.

    “Recent work showed that E1A can inhibit (11, 25, 57) or stimulate (1) CBP-AT activity.”
    Quote in the paperNot a substantial claim1 of 3 sources checked
    • cites [11] source
    • cites [25] source
    • cites [57] source

      Citation (1) is attached to the claim that E1A stimulates CBP acetyltransferase activity. The Results report that E1A markedly inhibited CBP-dependent histone acetylation; the paper instead identifies p/CIP as a factor that can stimulate CBP acetyltransferase activity (abstract; page not available in our copy). (LLM, conf=0.91)

      Checked by a reasoning model — may be wrong.

      Experimental — accuracy not yet measured

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

    Premisecitation
    § Introduction

    E1A blocks terminal differentiation of MEL cells, implicating CBP and p300 as critical cofactors for erythroid transcriptional regulators.

    “E1A blocks terminal differentiation of MEL cells, implicating CBP and p300 as critical cofactors for erythroid transcriptional regulators (9).”
    Quote in the paperSubstantial claim1 of 1 sources checked

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

    Premisecitation
    § Introduction

    CBP binds to GATA-1 and stimulates its transcriptional activity.

    “Our previous work showed that CBP binds to GATA-1 and stimulates its transcriptional activity (9).”
    Quote in the paperNot a substantial claim1 of 1 sources checked

  • May combine two claims.

    Premisecitation
    § Introduction

    GATA-1 is essential for differentiation and survival of erythroid precursor cells.

    “GATA-1 is essential for differentiation and survival of erythroid precursor cells (19, 79) and participates in the regulation of all erythroid-expressed genes tested to date (for a review see reference 78).”
    Quote in the paperSubstantial claim1 of 2 sources checked
    • cites [19] source
    • cites [79] source

  • May combine two claims.

    Premisecitation
    § Introduction

    Both E1A and PU.1 can block GATA-1 activity and cellular differentiation.

    “Both proteins can block GATA-1 activity and cellular differentiation.”
    Quote in the paperSubstantial claim2 of 3 sources checked
    • cites [9] source

      The Results report that wild-type E1A blocks CBP-stimulated GATA-1-dependent transcription, and the Discussion reports that activated E1A blocks differentiation in murine erythroleukemia cells. Our copy does not report that PU.1 blocks either GATA-1 activity or differentiation (searched for PU.1). No page numbers were supplied for these passages. (LLM, conf=0.88)

      Experimental — accuracy not yet measured
    • cites [40] source
    • cites [63] source

      The section on PU.1 domains reports that PU.1 represses GATA-1-mediated transcription and blocks differentiation in MEL cells. The passages found for E1A do not address whether E1A has either effect, so this source supports only the PU.1 part of the claim. (LLM, conf=0.85)

      Experimental — accuracy not yet measured

  • May combine two claims.

    Premisecitation
    § Introduction

    The CBP-binding domains of E1A and PU.1 are required for blocking erythroid differentiation and for inhibition of GATA-1 activity.

    “their respective CBP-binding domains are required for blocking erythroid differentiation and for inhibition of GATA-1 activity (9, 40, 63, 82).”
    Quote in the paperSubstantial claim2 of 3 sources checked
    • cites [9] source

      The Results (page not specified) show that wild-type E1A inhibits CBP-stimulated GATA-1 transcription, while a CBP-binding-defective mutant has little effect. They also report that E1A blocks erythroid differentiation in murine erythroleukemia cells, whereas CBP-binding-defective mutants do not. The cited paper does not establish the corresponding claim about PU.1. (LLM, conf=0.88)

      Experimental — accuracy not yet measured
    • cites [40] source
    • cites [63] source

      The sections on PU.1-mediated inhibition of GATA-1 activity and blocking erythroid differentiation report that PU.1’s transactivation and DNA-binding domains are required for both effects (page unavailable). The paper does not identify these as CBP-binding domains or establish that E1A’s CBP-binding domain is required for either effect. (LLM, conf=0.88)

      Experimental — accuracy not yet measured

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

    Premisecitation
    § Introduction

    GATA-1, EKLF, and NF-E2 interact with CBP, and their activities are inhibited by E1A.

    “Indeed, three erythroid-expressed transcription factors, GATA-1, erythroid Krüppel-like factor (EKLF), and NF-E2, which are important for erythroid differentiation and globin gene expression, interact with CBP, and their activities are inhibited by E1A (9, 14, 18, 28, 85).”
    Quote in the paperNot a substantial claim3 of 3 sources checked
    • cites [9] source

      The Abstract and Results show that GATA-1 associates with CBP and that E1A blocks CBP-enhanced GATA-1 transcription. The Discussion reports an in vitro CBP interaction for NF-E2’s p45 subunit but calls EKLF only a candidate for interaction; it does not establish that E1A inhibits NF-E2 or EKLF activity. (LLM, conf=0.91)

      Experimental — accuracy not yet measured
    • cites [14] source

      The Results and Discussion (page unavailable in our copy) show that p45/NF-E2 interacts with CBP. They do not establish CBP interactions for GATA-1 or EKLF, or show that E1A inhibits the activities of any of the three factors; the reported inhibition instead uses a CBP fragment. (LLM, conf=0.87)

      Experimental — accuracy not yet measured
    • cites [18] source

      The HS2 E1A-sensitivity section (page unavailable) says GATA-1 and NF-E2 interact with CBP/p300, and the Discussion (page unavailable) reports that E1A inhibits transactivation mediated by NF-E2’s p45 activation domain. The paper mentions EKLF as a reported substrate for CBP/p300 acetylation, but does not establish the claimed CBP interaction or E1A inhibition of EKLF activity. It also does not show that E1A inhibits GATA-1 activity itself. (LLM, conf=0.86)

      Experimental — accuracy not yet measured

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

    Premisecitation
    § Introduction

    CBP and p300 are targets of several viral oncoproteins.

    “CBP and its close relative p300 are targets of several viral oncoproteins, including adenovirus E1A, simian virus 40 T, human papillomavirus E6, Epstein-Barr virus Zta, and the Kaposi's sarcoma-associated herpesvirus protein viral interferon regulatory factor (for a review see reference 21).”
    Quote in the paperNot a substantial claim1 of 1 sources checked
    • cites [21] source

      The ‘Association of CBP/p300 with viral oncoproteins’ section identifies E1A, SV40 T antigen, and HPV E6 as proteins that affect CBP/p300. The ‘Other viruses’ section says Kaposi sarcoma-associated herpesvirus vIRF binds CBP/p300, but our copy does not identify the listed Epstein–Barr virus protein Zta; it discusses EBNA2 instead. Thus the general claim is supported, but not every example in the citing sentence is verified. (LLM, conf=0.84)

      Experimental — accuracy not yet measured

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

    Premisecitation
    § Introduction

    Expression of GATA-1 in multipotent progenitor cells triggers erythroid differentiation with a concomitant reduction in PU.1 expression and a block in myeloid differentiation.

    “Conversely, expression of GATA-1 in these cells triggers erythroid differentiation with a concomitant reduction in PU.1 expression and a block in myeloid differentiation (31, 42).”
    Quote in the paperNot a substantial claim1 of 2 sources checked
    • cites [31] source
    • cites [42] source

      The section ‘GATA-1: both an activator and a repressor of the EOS47 promoter’ (page unavailable) reports that activating GATA-1 in chicken myeloblasts rapidly down-regulates myeloid markers. It describes differentiation toward eosinophils at low GATA-1 levels, not erythroid differentiation of multipotent progenitors, and does not report a reduction in PU.1 expression or a block in myeloid differentiation. (LLM, conf=0.82)

      Experimental — accuracy not yet measured

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

    Premisecitation
    § Introduction

    CBP acetylates GATA-1 at two highly conserved lysine-rich motifs near each of the two zinc fingers.

    “CBP acetylates GATA-1 at two highly conserved lysine-rich motifs near each of the two zinc fingers (10, 27).”
    Quote in the paperNot a substantial claim1 of 2 sources checked
    • cites [10] source
    • cites [27] source

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

    Premisecitation
    § RESULTS

    CBP binds and acetylates EKLF.

    “Recent work showed that CBP binds and acetylates EKLF (85, 86).”
    Quote in the paperSubstantial claim1 of 2 sources checked
    • cites [85] source
    • cites [86] source

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

    Premisecitation
    § RESULTS

    p53 is acetylated by both PCAF and CBP, and acetylation strongly stimulates DNA binding to naked DNA in vitro.

    “p53 is acetylated by both PCAF and CBP, and acetylation strongly stimulates DNA binding to naked DNA in vitro (22, 36, 65).”
    Quote in the paperNot a substantial claim2 of 3 sources checked
    • cites [22] source
    • cites [36] source
    • cites [65] source

      The sections “PCAF and p300 acetylate p53” and “Acetylation by p300 and PCAF activate sequence-specific DNA-binding” report that both enzymes acetylated p53 and that acetylation strongly increased its binding to a synthetic DNA probe in vitro. The experiments used p300, not CBP; although the paper describes CBP and p300 as closely related, it does not establish the claim specifically for CBP. Page numbers are unavailable in our copy. (LLM, conf=0.87)

      Experimental — accuracy not yet measured

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

    Premisecitation
    § DISCUSSION

    The viral interferon regulatory factor of Kaposi's sarcoma-associated herpesvirus has transforming activity and inhibits p300-mediated histone acetylation.

    “An ex- ample to support this hypothesis is provided by the viral inter- feron regulatory factor of Kaposi's sarcoma-associated herpes- virus, which has transforming activity and has been shown to inhibit p300-mediated histone acetylation (34).”
    Quote in the paperNot a substantial claim1 of 1 sources checked

  • May combine two claims.

    Premisecitation
    § DISCUSSION

    Loss of GATA-1 function not only results in a differentiation block but also leads to apoptotic cell death.

    “However, it is important to note that loss of GATA-1 function not only results in a differentiation block but also leads to apoptotic cell death (79).”
    Quote in the paperSubstantial claim1 of 1 sources checked

  • Classified as probably not a substantial claim.

    Premisecitation
    § DISCUSSION

    Acetylation of GATA-1 is important for GATA-1 function in differentiating erythroid cells.

    “Previous work has indicated that acetylation of GATA-1 is important for GATA-1 function in differentiating erythroid cells (27).”
    Quote in the paperSubstantial claim1 of 1 sources checked

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

    Premisecitation
    § DISCUSSION

    The cellular protein E1A-like inhibitor of differentiation (EID-1) can inhibit CBP/p300-AT activity and block myogenic differentiation.

    “A recently identified cellular protein called E1A-like inhibitor of differentiation (EID-1) can inhibit CBP/ p300-AT activity and block myogenic differentiation (37, 47).”
    Quote in the paperNot a substantial claim1 of 2 sources checked
    • cites [37] source
    • cites [47] source

  • May combine two claims.

    Premisecitation
    § DISCUSSION

    In both yeast and mammalian cells, histone acetylation precedes transcription and occurs even in the absence of transcription.

    “Both in yeast and in mam- malian cells histone acetylation precedes transcription and occurs even in the absence of transcription (17, 26, 30, 33, 67).”
    Quote in the paperSubstantial claim3 of 3 sources checked
    • cites [17] source

      The Results and Discussion report an inactive murine βminor promoter that was hyperacetylated and show that increasing acetylation at the Ey promoter did not induce transcription. These findings support acetylation without transcription in the mammalian system studied. The paper does not establish that acetylation temporally precedes transcription or report a corresponding finding in yeast; page numbers were unavailable in our copy. (LLM, conf=0.86)

      Experimental — accuracy not yet measured
    • cites [26] source

      The Analysis of immunofractionated chromatin section (page not provided) reports acetylated histones on chicken globin genes even when those genes are not transcribed. The Discussion links acetylation to transcriptional competence, but the paper does not establish that acetylation precedes transcription in time or demonstrate the claim in both yeast and mammalian cells. (LLM, conf=0.88)

      Experimental — accuracy not yet measured
    • cites [30] source

      The section on nucleosomes encompassing the HO promoter reports that acetylation precedes HO transcription by about 15–30 minutes. The section on histone H3 acetylation at the HO promoter reports acetylation in a yeast swi4 mutant with no HO transcription. These passages support the yeast finding, but the paper does not establish the same sequence or transcription-independent acetylation in mammalian cells. (LLM, conf=0.88)

      Experimental — accuracy not yet measured

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

    Premisecitation
    § DISCUSSION

    The cellular protein Twist can inhibit p300-AT and PCAF-AT activities and has oncogenic potential.

    “In addition, the cellular protein Twist, which can inhibit p300-AT and PCAF-AT activities (25), has been shown to have oncogenic potential (38).”
    Quote in the paperNot a substantial claim1 of 2 sources checked
    • cites [25] source
    • cites [38] source

  • May combine two claims.

    Findingown data
    § RESULTS

    While histone H3 acetylation increased approximately twofold in control MEL cells, no change in histone H3 acetylation was observed in both MEL-PU.1 clones at HS3.

    “While histone H3 acetylation increased approximately twofold in control MEL cells, no change in histone H3 acetylation was observed in both MEL-PU.1 clones (Fig. 11B).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data
    § Introduction

    PU.1 inhibits CBP-mediated acetylation of GATA-1 in vitro and in vivo.

    “Here we demonstrate that PU.1 inhibits CBP-mediated acetylation of GATA-1 in vitro and in vivo.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data
    § RESULTS

    Upon coexpression of PU.1, reporter activity was dramatically reduced, whereas the basal activity of the Bax promoter was not inhibited.

    “Upon coexpression of PU.1, reporter activity was dramatically reduced, whereas the basal activity of the Bax promoter was not inhibited (Fig. 7C).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data
    § DISCUSSION

    Inhibition of PCAF required higher PU.1 concentrations and was less complete than the inhibition of CBP, suggesting a degree of specificity.

    “However, inhibition of PCAF required higher PU.1 concentrations and was less complete than the inhibition of CBP, suggesting a degree of specificity.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data
    § DISCUSSION

    GATA-1, EKLF, MafG, and p53 do not all inhibit histone acetylation, contrary to what would be expected if PU.1 inhibited their acetylation simply by competing for a single binding site within CBP.

    “However, if PU.1 were to inhibit GATA-1, EKLF, MafG, and p53 acetylation simply by competing for a single binding site within CBP, all these factors would consequently be ex- pected to inhibit histone acetylation. However, our results show that this is not the case.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingnone given
    § DISCUSSION

    PU.1 inhibits acetylation and activity of the tumor suppressor protein p53.

    “In this context it is noteworthy that PU.1 inhibits acetylation and activity of the tumor sup- pressor protein p53.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based).

    Findingown data
    § DISCUSSION

    PU.1 competes with GATA-1 and NF-E2 for binding to the CH3 domain of CBP.

    “Preliminary protein-binding studies showed that PU.1 com- petes with GATA-1 and NF-E2 for binding to the CH3 domain of CBP (unpublished observation).”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingnone given
    § DISCUSSION

    PU.1 inhibited acetylation and transcriptional activity of the hematopoietic transcription factors NF-E2 and EKLF.

    “Indeed, PU.1 inhibited acetylation and transcriptional activity of the hematopoietic transcription factors NF-E2 and EKLF.”
    Quote in the paperSubstantial claim

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

    Premisecitation
    § Introduction

    PU.1 is essential for the formation of the myeloid and lymphoid cell lineages.

    “In the normal hematopoietic system, PU.1 is essential for the formation of the myeloid and lymphoid cell lineages (41, 71; for reviews see references 16 and 49).”
    Quote in the paperNot a substantial claim0 of 3 sources checked
    • cites [41] source
    • cites [71] source
    • cites [16] source

  • May combine two claims.

    Premisecitation
    § Abstract

    Aberrant PU.1 expression in erythroid precursor cells inhibits erythroid differentiation and causes murine erythroleukemias.

    “Aberrant expression of PU.1 inhibits erythroid cell differentiation and contributes to the formation of murine erythroleukemias (MEL).”
    Quote in the paperSubstantial claim0 of 3 sources checked
    • cites [50] source
    • cites [16] source
    • cites [49] source
    • Rests on an assumption: The molecular mechanism by which aberrant PU.1 expression inhibits erythroid differentiation is poorly understood.

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

    Interpretationcitation
    § Introduction

    GATA-1 can directly inhibit PU.1 activity.

    “suggesting that GATA-1 can directly inhibit PU.1 activity (54; see below).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [54] source

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

    Premisecitation
    § RESULTS

    Histone acetylation is an early event during gene activation and sets the stage for subsequent transcriptional events.

    “Thus, it is believed that histone acetylation is an early event during gene activation and sets the stage for subsequent transcriptional events.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Premisecitation
    § RESULTS

    The tethered NF-E2 construct is fully active and can restore NF-E2 activity in NF-E2-deficient erythroid cells.

    “This construct is fully active and can restore NF-E2 activity in NF-E2-deficient erythroid cells (7).”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [7] source

  • May combine two claims.

    Interpretationown data
    § RESULTS

    PU.1 may block erythroid differentiation by targeting not only GATA-1 but also multiple CBP-dependent erythroid transcription factors.

    “Together, these results suggest that PU.1 may block erythroid differentiation by targeting not only GATA-1 but also multiple CBP-dependent erythroid transcription factors.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationown data
    § RESULTS

    PU.1 action is specific toward certain substrates and PU.1 does not inhibit the AT activity of CBP per se but might instead regulate accessibility of CBP substrates.

    “suggesting that PU.1 action is specific toward certain substrates and that PU.1 does not inhibit the AT activity of CBP per se but might instead regulate accessibility of CBP substrates.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Premiseown data
    § DISCUSSION

    Both E1A and PU.1 bind to the CH3 domain of CBP but require additional contacts for full CBP inhibition.

    “Both E1A and PU.1 bind to the CH3 domain of CBP but require additional contacts for full CBP inhibition.”
    Quote in the paperSubstantial claim

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

    Interpretationcitation
    § DISCUSSION

    The discrepancy between these reports might be the result of differences in E1A dosage where inhibition of CBP-AT activity requires high concentrations of E1A.

    “The discrepancy between these reports might be the result of differences in E1A dosage where inhibition of CBP-AT activity requires high concentrations of E1A (35).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [35] source

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

    Premisecitation
    § DISCUSSION

    The mutually repressive effects between c-Myb and GATA-1 were attributed to the failure of these proteins to simultaneously bind to CBP.

    “Similarly, the mutually re- pressive effects between c-Myb and GATA-1 were attributed to the failure of these proteins to simultaneously bind to CBP (74).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [74] source

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

    Interpretationreasoning
    § DISCUSSION

    The potency of acetylation inhibition by PU.1 might be determined by the relative affinities of PU.1 and acetylation substrates for the AT domain of CBP.

    “Another prediction from this mech- anism would be that the potency of acetylation inhibition by PU.1 might be determined by the relative affinities of PU.1 and acetylation substrates for the AT domain of CBP.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S1 (asserted), S3 (load-bearing).

    Interpretationnone given
    § DISCUSSION

    Inhibition of GATA-1 DNA binding requires very high concentrations of PU.1.

    “It is possible that inhibition of GATA-1 DNA binding requires very high concentrations of PU.1.”
    Quote in the paperNot a substantial claim

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

    Interpretationreasoning
    § DISCUSSION

    PU.1 may interact with substrate proteins to prevent their interaction with CBP.

    “It is also possible that PU.1 interacts with substrate proteins to prevent their interaction with CBP.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Interpretationreasoning
    § DISCUSSION

    The results are consistent with a model in which inhibition of histone acetylation by PU.1 is the primary event, followed by inhibition of gene expression and cellular differentiation.

    “Thus, our results are consistent with a model in which inhibi- tion of histone acetylation by PU.1 is the primary event, fol- lowed by inhibition of gene expression and cellular differenti- ation.”
    Quote in the paperSubstantial claim

  • Classified as probably not a substantial claim.

    Interpretationreasoning
    § DISCUSSION

    The high efficiency of PU.1 in inhibiting GATA-1 acetylation may be the result of PU.1's ability to bind to both CBP and GATA-1.

    “Thus, it is possi- ble that the high efficiency of PU.1 in inhibiting GATA-1 acetylation is the result of PU.1's ability to bind to both CBP and GATA-1.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Premisecitation
    § DISCUSSION

    Both E1A and PU.1 bind RB.

    “Of note, both E1A and PU.1 bind RB, further extending their functional similarities (24).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [24] source

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

    Premisecitation
    § DISCUSSION

    PU.1 activates numerous myeloid and lymphoid tissue-expressed genes.

    “PU.1 acti- vates numerous myeloid and lymphoid tissue-expressed genes (16, 49)”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [16] source
    • cites [49] source

  • Classified as probably not a substantial claim.

    Premisecitation
    § DISCUSSION

    CBP stimulates PU.1 activity in transient transfections.

    “and CBP stimulates PU.1 activity in transient trans- fections (82).”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [82] source

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

    Premisecitation
    § DISCUSSION

    The progression of neurodegeneration is slowed by treatment with deacetylase inhibitors.

    “The progression of neurodegeneration is slowed by treatment with deacetylase inhibitors, further emphasizing the impor- tance of a controlled balance of protein acetylation and deacetylation in cellular functions in diverse tissues (73).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [73] source

  • May combine two claims.

    Interpretationreasoning
    § DISCUSSION

    The presence or absence of PU.1-binding sites, as well as promoter architecture, might determine whether PU.1 activates or inhibits transcription of a given gene.

    “Thus, the presence or absence of PU.1-binding sites, as well as promoter architecture, might determine whether PU.1 activates or inhibits transcription of a given gene.”
    Quote in the paperSubstantial claim

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

    Interpretationreasoning
    § DISCUSSION

    At high concentrations, PU.1 might be able to occupy and activate myeloid and lymphoid promoters while leaving enough PU.1 protein available to inhibit the activities of erythroid transcription factors.

    “At high concentrations PU.1 might be able to occupy and activate myeloid and lymphoid promoters while leaving enough PU.1 protein available to inhibit the activities of erythroid transcrip- tion factors.”
    Quote in the paperSubstantial claim

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

    Premisecitation
    § DISCUSSION

    Low concentrations of PU.1 in hematopoietic precursor cells are permissive for a developmental path toward the erythroid cell fate, while higher levels favor differentiation toward the myeloid and lymphoid lineage and inhibit erythroid differentiation.

    “Previous studies showed that low concentra- tions of PU.1 in hematopoietic precursor cells are permissive for a developmental path toward the erythroid cell fate, while higher levels favor differentiation toward the myeloid and lym- phoid lineage and inhibit erythroid differentiation (see the introduction).”
    Quote in the paperNot a substantial claim

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

    Premisecitation
    § DISCUSSION

    PU.1 levels are critical in determining myeloid versus lymphoid cell fate.

    “Furthermore, PU.1 levels appear to be critical in determining myeloid versus lymphoid cell fate (15).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [15] source

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

    Premisecitation
    § DISCUSSION

    Neurodegeneration triggered by expression of the polyglutamine-containing protein Httex1p involves inhibition of CBP/p300- and PCAF-AT activities.

    “A recent report showed that neurodegeneration triggered by expression of the polyglutamine-containing protein Httex1p involves inhibition of CBP/p300- and PCAF-AT activities (73).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [73] source

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

    Premisecitation
    § DISCUSSION

    In the normal hematopoietic system, PU.1 is essential for myeloid and lymphoid cell development.

    “In the normal hematopoietic system PU.1 is essential for myeloid and lymphoid cell development (41, 71).”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [41] source
    • cites [71] source

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

    Premisecitation
    § DISCUSSION

    Components of the INHAT complex can inhibit p300-mediated histone acetylation and are potentially oncogenic.

    “Also, components of the INHAT complex, which can inhibit p300-mediated histone acetylation, are potentially oncogenic (72).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [72] source
52 of 52

  • The paper relies on this without support.

    The molecular mechanism by which aberrant PU.1 expression inhibits erythroid differentiation is poorly understood.

    “The molecular mechanism by which this occurs is poorly understood.”

  • The paper relies on this without support.

    GAL4-CBP-AT consists largely of the AT domain of CBP.

    “Since GAL4-CBP-AT consists largely of the AT domain of CBP, this further suggests a direct effect of PU.1 on CBP function.”

  • The paper relies on this without support.

    Various transformed cell lines of diverse cellular origins differentiate in response to deacetylase inhibitors.

    “Since various transformed lines of diverse cellular origins differenti- ate in response to deacetylase inhibitors”
3 of 3

Claims other papers make about this one

Each is checked against this paper’s own text: does it say what the citing paper says it does?

  • CBP acetyltransferase activity increases during MEL cell differentiation as PU.1 levels decline and is inhibited by sustained PU.1 expression, suggesting importance of a regulated balance between CBP and PU.1 during erythroid cell differentiation.

    “Furthermore, CBP acetyltransferase activity has been reported to increase during MEL cell differentiation as PU.1 levels decline and is inhibited by sustained PU.1 expression [81], suggesting the importance of a regulated balance between CBP and PU.1 during erythroid cell differentiation.”

    From PU.1 and partners: regulation of haematopoietic stem cell fate in normal and malignant haematopoiesis (2009) · cites this paper as [81]

  • During differentiation of murine erythroleukaemia (MEL) cells, there is an inverse correlation between PU.1 levels and CBP-acetyl transferase activity.

    “During differentiation of murine erythroleukaemia (MEL) cells there is an inverse correlation between PU.1 levels and CBP-acetyl transferase activity [81].”

    From PU.1 and partners: regulation of haematopoietic stem cell fate in normal and malignant haematopoiesis (2009) · cites this paper as [81]

  • In MEL cells, Spi-1 alters the transcriptional activities of GATA-1 by interacting with GATA-1 bound on DNA and by repressing GATA-1 target genes.

    “In the MEL cells, Spi-1 alters the transcriptional activities of GATA-1 by interacting with GATA-1 bound on DNA and by repressing GATA-1 target genes (20–22,39).”

    From Spi-1/PU.1 activates transcription through clustered DNA occupancy in erythroleukemia (2012) · cites this paper as [20]

Self-check

Percentages, sums and reported differences re-computed from the paper’s own numbers.

Each contradiction between two passages listed here was confirmed by an independent check.

  • PU.1 deletion construct has conflicting boundaries

    Minor

    The construct described as lacking most of the activation domain is named PU.1�30-100, but the subsequent results name it PU.1�33-100. The text does not establish that these are separate constructs.

1 of 1

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Authors
Hong, W., Kim, A. Y., Ky, S., Rakowski, C., Seo, S.-B., Chakravarti, D., Atchison, M., & Blobel, G. A.
Affiliations
  • Division of Hematology, Children's Hospital of Philadelphia
  • Children's Hospital of Philadelphia (US)
  • Department of Pharmacology
  • University of Pennsylvania (US)
Corresponding author
Blobel, Gerd A.
ORCID
Venue
Molecular and Cellular Biology
Publisher
Informa UK Limited
Published
2002-06-01
Volume
22
Issue
11
Pages
3729-3743
ISSN
  • 0270-7306 (ISSN-L)
  • 1098-5549
DOI
10.1128/mcb.22.11.3729-3743.2002
PMC
PMC133832
OpenAlex
W1980156635
Cited by
71
References
91
Citations by year
2026: 1 · 2025: 3 · 2022: 1 · 2021: 2 · 2020: 2 · 2019: 1 · 2018: 3 · 2014: 2 · 2013: 1 · 2012: 4
FWCI
3.16
Type
journal article
Language
en
Open access
Licence
journals.asm.org/non-commercial-tdm-license (tdm)
Locations
  • Molecular and Cellular Biology (publishedVersion)
  • PubMed (publishedVersion)
  • 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 › Epigenetics and DNA Methylation
Keywords
PU.1, CBP, protein acetylation, erythroid differentiation, GATA-1, NF-E2, Krüppel-like factors, histone acetylation
MeSH terms
Proto-Oncogene Protein Spi-1, Acetylation, Animals, Cell Differentiation, Cell Division, Cell Line

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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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