HMGA1 chromatin regulators drive transcriptional networks involved in megakaryocyte expansion, fibrosis, and sensitivity to interferon signaling in JAK2-V617F MPN

Abstract Introduction: Myeloproliferative neoplasms (MPN) are clonal hematopoietic stem cell (HSC) disorders with an increased risk of transformation to secondary myelofibrosis (sMF) or acute leukemia. Although clinical outcomes are poor after transformation, actionable mechanisms that drive progression remain elusive. The gene encoding the High Mobility Group A1 chromatin regulator is overexpressed with MPN progression and required for leukemic transformation in preclinical models (Li et al, Blood 2022). Unexpectedly, loss of just a single Hmga1 allele within the HSC compartment also decreases thrombocytosis and erythrocytosis while preventing splenomegaly and progression to sMF in JAK2V617F mice. Intriguingly, HMGA1 also drives fibrosis and progression in diverse tumors (Chia et al, JCI 2023; Luo et al, JCI 2025). Multiomic studies reveal that HMGA1 modulates pro-fibrotic and inflammatory pathways, including IFN signaling, suggesting that HMGA1 may impact sensitivity to IFNα therapy (IFNα). We therefore sought to: 1) Define targetable mechanisms underlying HMGA1 early in MPN progression, 2) Elucidate the role of HMGA1 in sensitivity to IFNα. Methods: To elucidate mechanisms underlying HMGA1 early in MPN, we compared transcriptomes via single cell RNA sequencing (scRNAseq) in HSC and progenitor cells (HSPCs) from bone marrow of wildtype (WT) or JAK2V617F mice with varied levels of Hmga1. To assess responses to IFNα, we compared MPN phenotypes and single cell transcriptomes in HSPC. We also queried transcriptomes from patient peripheral blood mononuclear cells (PBMCs), platelets, and CD34+ cells. Gene set enrichment analysis (GSEA) was used to define cellular pathways. Results: Hmga1 haploinsufficiency within HSCs dampens expansion of long-term, quiescent HSC (qHSC), megakaryocyte-biased HSC (Mk-HSC), and megakaryocyte-erythroid-biased HSC (MEP-HSC) while expanding lymphoid biased HSC (Ly-HSC) in JAK2V617Fmice with MPN prior to developing sMF. Trajectory analysis reveals that JAK2V617F HSC with intact Hmga1 have decreased differentiation capacity compared to those with Hmga1 heterozygosity. Because Hmga1 drives megakaryocyte hyperplasia and thrombocytosis in JAK2V617Fmice, we identified Hmga1 gene networks within Mk-HSC and MEP-HSC, which show that Hmga1 activates genes governing: 1) cell cycle progression (MYC Targets, E2F Targets, Mitotic Spindle), 2) metabolism required for HSC cycling oxidative phosphorylation (OxPhos), 3) coagulation and platelet activation platelet factor 4, von Willebrand factor, glycoprotein (Gp) 2b, Gp3a, P-selectin. Unexpectedly, Hmga1 also induces IFNα signaling, including IFNα receptor and downstream effector genes, suggesting that HMGA1 may enhance sensitivity to IFNα in JAK2V617F HSPC. In JAK2V617F mice with intact Hmga1, IFNα results in normal blood counts and spleen sizes. IFNα also decreases mutant qHSC and granulocyte-monocyte progenitors (GMPs), while increasing the proportion of cycling HSC and multipotent progenitors (cycHSC/MPP). By contrast, in WT mice, IFNα has no impact on blood counts nor spleen size with minimal effects on qHSC. However, IFNα pushes WT cycHSC/MPP to more committed GMP and GSEA reveals that IFNα therapy activates proliferation pathways in WT HSPC. In JAK2V617F HSPC, IFNα activates IFN response genes while repressing complement, TNFα-NF-κB, and coagulation pathways, suggesting that IFNα therapy preferentially decreases a subset of inflammatory and pro-thrombotic gene networks in JAK2V617F cells. Patient MPN samples (platelets, PBMCs, CD34+cells) show activation of similar HMGA1 transcriptional pathways, highlighting the relevance of our results from JAK2V617F mouse models in humans. Conclusions: We uncovered a novel epigenetic program in JAK2V617FMPN whereby Hmga1 induces gene networks that drive expansion in mutant HSPC with greatest impact on HSC and progenitors poised to differentiate into megakaryocytes, granulocytes, and monocytes (Mk-HSC, MEP-HSC, GMP). Mechanistically, Hmga1 up-regulates genes involved in platelet activation, inflammation, and IFN signaling in JAK2V617F HSPC and these pathways are preferentially targeted by IFNα therapy in JAK2V617F HSPC compared to WT HSPC. Together, our studies reveal a new paradigm whereby HMGA1 drives progression early in MPN by activating gene networks required for megakaryocyte expansion, fibrosis, inflammatory signaling, and IFN networks, the latter of which may sensitize JAK2 mutant HSPC to IFNα therapy.