ETV6 germline mutations cause HDAC3/NCOR2 mislocalization and upregulation of interferon response genes

Fisher, Marlie H; Kirkpatrick, Gregory D.; Stevens, Brett M.; Jones, Courtney L.; Callaghan, Michael U.; Rajpurkar, Madhvi; Fulbright, Joy M.; Cooper, Megan A.; Rowley, Jesse W.; Porter, Christopher C.; Gutierrez‐Hartmann, Arthur; Jones, Kenneth L.; Jordan, Craig T.; Pietras, Eric M.; Paola, Jorge Di

doi:10.1172/jci.insight.140332

Cited by 18 publications

(14 citation statements)

References 62 publications

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“…2i,j and Extended Data Fig. 3e ), consistent with a previous report 24 . These results show that ETV3 and ETV6 repress ISG expression in monocytes in vitro and in vivo in humans and suggest that STAT1 signaling may be involved in the differentiation of monocytes.…”

Section: Resultssupporting

confidence: 92%

“…2f,g ). To determine the in vivo relevance of this finding, we reanalyzed scRNA-seq data from peripheral blood mononuclear cells (PBMCs) of patients carrying a germline mutation of ETV6 (P214L) resulting in loss of function 24 . We first filtered the data to retain only CD14 + and CD16 + monocytes from healthy and ETV6 P214L patients (Extended Data Fig.…”

Section: Resultsmentioning

confidence: 99%

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ETV3 and ETV6 enable monocyte differentiation into dendritic cells by repressing macrophage fate commitment

et al. 2022

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In inflamed tissues, monocytes differentiate into macrophages (mo-Macs) or dendritic cells (mo-DCs). In chronic nonresolving inflammation, mo-DCs are major drivers of pathogenic events. Manipulating monocyte differentiation would therefore be an attractive therapeutic strategy. However, how the balance of mo-DC versus mo-Mac fate commitment is regulated is not clear. In the present study, we show that the transcriptional repressors ETV3 and ETV6 control human monocyte differentiation into mo-DCs. ETV3 and ETV6 inhibit interferon (IFN)-stimulated genes; however, their action on monocyte differentiation is independent of IFN signaling. Instead, we find that ETV3 and ETV6 directly repress mo-Mac development by controlling MAFB expression. Mice deficient for Etv6 in monocytes have spontaneous expression of IFN-stimulated genes, confirming that Etv6 regulates IFN responses in vivo. Furthermore, these mice have impaired mo-DC differentiation during inflammation and reduced pathology in an experimental autoimmune encephalomyelitis model. These findings provide information about the molecular control of monocyte fate decision and identify ETV6 as a therapeutic target to redirect monocyte differentiation in inflammatory disorders.

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Section: Resultssupporting

confidence: 92%

Section: Resultsmentioning

confidence: 99%

ETV3 and ETV6 enable monocyte differentiation into dendritic cells by repressing macrophage fate commitment

et al. 2022

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“…The copyright holder for this preprint this version posted December 11, 2022. ; https://doi.org/10.1101/2022.12.08.519689 doi: bioRxiv preprint All MPN patients in the cohort were recruited at the Hematology Department of Washington University School of Medicine in St. Louis. Sample processing were performed as previously published 41 . Chromium Next GEM Single cell 3′ Reagent v3.1 (Dual Index) was used for GEM Generation, barcoding and cDNA library preparation per manufacturer's guidelines, and cDNA was sequenced using the 10X Genomics Single-cell RNA-Seq platform at the Genome Technology Access Center at the McDonnell Genome Institute of Washington University in St. Louis.…”

Section: Platelet Aggregation Assaymentioning

confidence: 99%

Multiomic Profiling Reveals Metabolic Alterations Mediating Aberrant Platelet Activity and Inflammation in Myeloproliferative Neoplasms

Laranjeira

Kong

et al. 2022

Preprint

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Prior studies by our group and others have demonstrated that platelets from patients with myeloproliferative neoplasms (MPNs) exhibit a hyperreactive phenotype. However, a complete understanding of platelet alterations in MPNs remains lacking, and the mechanisms by which platelets contribute to MPN-related thrombosis, as well as other MPN disease features, are incompletely understood. In this study, utilizing multiomic approaches to interrogate platelet phenotypes in patient samples, in conjunction with relevant MPN animal models, we aim to investigate mechanisms of dysregulated platelet activity in MPNs, and explore how these findings may be leveraged to uncover novel therapeutic strategies. We initially studied platelet activation in peripheral blood from patients with essential thromobocythemia (ET) and compared to age-, sex-matched healthy controls. We found significantly increased P-selectin exposure in conjunction with increased platelet-leukocyte aggregates indicating activation of platelets from ET patients. To investigate the transcriptional signature of PBMCs and platelets at the single cell level, we performed single cell RNA-seq (scRNA-seq) in PBMCs from ET patients and healthy controls. Monocytes were increased in ET patients and displayed the highest inflammation index, implicating monocytes as the primary source of inflammation in ET. Aside from the quantitative increases of platelets in ET patients, we demonstrated strong enrichment of platelet activation, mTOR and oxidative phosphorylation (OXPHOS) genes in platelets from ET patients. Proteomic data confirmed the activation of OXPHOS and mTOR pathways in platelets from MPN patients. Metabolomics analysis revealed distinct metabolic phenotypes consisting of elevated tricarboxylic acid (TCA) cycle components, ATP generation and lower alpha-ketoglutarate (alpha-KG), in platelets from MPN patients, all consistent with enhanced OXPHOS and mTOR activities. Enhanced mitochondrial respiration at both baseline and after ex vivo stimulation with the platelet agonist thrombin receptor activator peptide (TRAP6) by extracellular flux analysis confirmed the bioenergetic alterations in platelets from MPN patients. alpha-KG is a key TCA cycle intermediate, which inhibits PI3K/AKT/mTOR pathway signaling via suppression of ATP synthase. alpha-KG supplementation drastically reduced oxygen consumption and ATP generation in platelets from MPN patients. We further investigated the effects of alpha-KG on MPN platelet activation. Ex vivo incubation of platelets from both MPN patients and Jak2 V617F knock-in mice with alpha-KG significantly reduced platelet surface P-selectin and integrin gpIIb/IIIa activation. Additionally, alpha-KG inhibited the spreading and adhesion of platelets from Jak2 V617F knock-in mice to fibrinogen-coated surfaces. Platelet phosphoblots demonstrated significant downregulation of p-AKT and p-ERK after treatment with alpha-KG, suggesting the involvement of PI3K/AKT/mTOR and MAPK pathways in the inhibitory effects of alpha-KG on platelet activation. Thus, alpha-KG inhibited platelet hyperreactivities in both human and mouse MPN samples. To test the therapeutic impact of alpha-KG on MPN disease features, we treated Jak2 V617F knock-in mice with alpha-KG for 6 weeks. Oral alpha-KG supplementation decreased splenomegaly and reduced elevated platelets and hematocrit. Additionally, monocytes were significantly decreased as early as 2 weeks after alpha-KG treatment in Jak2 V617F knock-in mice. Consistently, RNA-seq of bone marrow samples from alpha-KG treated mice revealed inhibition of heme metabolism, OXPHOS and mTOR signaling pathways. In ex vivo studies with MPN patient CD34+ cells, alpha-KG treatment for 10 days led to a decrease in CD41+ CD61+ cells, suggesting decreased megakaryocyte commitment. We further observed that alpha-KG incubation significantly decreased the secretion of proinflammatory cytokines from sorted CD14+ human monocytes. Mass cytometry analysis of whole blood from MPN patients demonstrated inhibition of MAPK pathway signaling after alpha-KG treatment. Taken together, these results suggest that alpha-KG supplementation may exert therapeutic effects through both direct inhibition of MPN platelet activity and via quenching of monocyte hyper-inflammation. In summary, these studies reveal a previously unrecognized metabolic disorder in platelets from MPN patients and highlight a prominent role for alpha-KG and mTOR signaling in aberrant MPN platelet activity and monocyte-driven inflammation. These findings have potential relevance for novel therapeutic approaches for MPN patients.

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“…repressor complex, which is also reflected in patient-derived platelet transcripts and promotes aberrant proplatelet formation in megakaryocytes. 42 To our knowledge, the function of HDAC3 in ITP has not yet been reported. Chidamide is an HDAC inhibitor that selectively acts on multiple HDAC subtypes, including HDAC3.…”

Section: Discussionmentioning

confidence: 93%

“…Moreover, Thpo is required for megakaryopoiesis in the early development of megakaryocyte precursors 40 . ETV6 is an essential transcription factor in hematopoiesis and megakaryocyte development, 41 and overexpression of HDAC3‐regulated interferon response genes may be led by abnormal cytoplasmic localization of ETV6 and the HDAC3/NCOR2 repressor complex, which is also reflected in patient‐derived platelet transcripts and promotes aberrant proplatelet formation in megakaryocytes 42 . To our knowledge, the function of HDAC3 in ITP has not yet been reported.…”

Section: Discussionmentioning

confidence: 99%

HDAC3 single‐nucleotide polymorphism rs2530223 is associated with increased susceptibility and severity of primary immune thrombocytopenia

Liu

Wang

Zhang

et al. 2022

Int J Lab Hematology

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Introduction: Primary immune thrombocytopenia (ITP) is an autoimmune hemorrhagic disorder characterized by a low platelet count and increased risk of bleeding.We previously reported that low-dose chidamide, a histone deacetylase (HDAC) inhibitor, restores immune tolerance in patients with ITP. This study aimed to evaluate the association of a single-nucleotide polymorphism (SNP) rs2530223 in the HDAC3 gene with susceptibility to ITP and its clinical features.Methods: Patients with ITP and age-matched healthy participants were recruited for this case-control study. Genotyping of the HDAC3 rs2530223 polymorphism was performed using MassARRAY platform.Results: Individuals with T allele of HDAC3 rs2530223 exhibited a 1.472-fold increased risk of ITP susceptibility (OR 1.472; 95% CI 1.100-1.969; p = .009), while ones with the TT genotype under the codominant and recessive models, and the TC/TT genotypes under the dominant model all revealed increased risk of ITP susceptibility (dominant odds ratio[OR] 1.965; 95% CI: 1.046-3.656; p = .036; codominant OR 2.264; 95% CI 1.175-4.360; p = .015; and recessive OR 1.512; 95% CI 1.028-2.224; p = .036, respectively). Regarding platelet counts in ITP patients, we observed that the TC/TT genotypes exhibited a 3.932-fold increased risk for platelet (PLT) <30 Â 10 9 /L (OR 3.932; 95% CI 1.426-10.842; p = .008). Conclusion:This study indicates that HDAC3 rs2530223 may be an important genetic factor related to ITP susceptibility and platelet count in ITP patients, providing new perspectives on disease progression, new therapeutic targets, and severity prediction.

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ETV6 germline mutations cause HDAC3/NCOR2 mislocalization and upregulation of interferon response genes

Cited by 18 publications

References 62 publications

ETV3 and ETV6 enable monocyte differentiation into dendritic cells by repressing macrophage fate commitment

ETV3 and ETV6 enable monocyte differentiation into dendritic cells by repressing macrophage fate commitment

Multiomic Profiling Reveals Metabolic Alterations Mediating Aberrant Platelet Activity and Inflammation in Myeloproliferative Neoplasms

HDAC3 single‐nucleotide polymorphism rs2530223 is associated with increased susceptibility and severity of primary immune thrombocytopenia

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