Effect of immune cytokines on megakaryocytopoiesis and platelet homeostasis : role of interleukin 21

La mégacaryocytopoïèse est le processus de différenciation des cellules souches hématopoïétiques en mégacaryocytes produisant les plaquettes sanguines. Des données de transcriptome indiquent la présence des récepteurs aux interleukines (IL) -10, -17A et -21 sur les mégacaryocytes. Mon travail montre que les IL-10 et -17 n’ont pas d’effet apparent sur la prolifération et la différenciation in vitro des mégacaryocytes à partir des progéniteurs hématopoïétiques humains CD34+. Cependant, l’IL-21 augmente la prolifération des progéniteurs mégacaryocytaires dérivés de ces cellules et sans modifier leur différenciation. Les mégacaryocytes de la moelle osseuse humaine expriment aussi le récepteur à l’IL-21, ce qui suggère un rôle direct de l’IL-21 sur la mégacaryocytopoïèse in vivo. De façon concordante, l’expression de l’IL-21 chez la souris stimule la mégacaryocytopoïèse ainsi que la production des plaquettes, mais augmente la clairance des plaquettes par les macrophages. Ces travaux suggèrent que durant les réponses immunitaires, l’expression de l’IL-21 par les lymphocytes T CD4+ activés module l’homéostasie des plaquettes sanguines.Megakaryocytopoiesis is the process by which hematopoietic stem cells give rise to megakaryocytes which in turn produce blood platelets. Transcriptional studies indicate the presence of interleukin (IL) -10, -17 and -21 receptors on megakaryocytes. This work shows that IL-10 and -17 have no apparent effect on in vitro proliferation and differentiation of megakaryocytes derived from human CD34+ hematopoietic progenitors. However, IL-21 increases the proliferation of megakaryocyte progenitors derived from these CD34+ cells without modifying their differentiation. Moreover, human bone marrow megakaryocytes express IL-21 receptor, suggesting a direct role of IL-21 on megakaryocytopoiesis in vivo. Concordantly, IL-21 expression in mice stimulates megakaryocytopoiesis and platelet production, but increases platelet clearance by macrophages. This work suggest that during immune responses, the expression of IL-21 by activated CD4+ T lymphocytes modulate blood platelet homeostasis

Effect of immune cytokines on megakaryocytopoiesis and platelet homeostasis : role of interleukin 21

La mégacaryocytopoïèse est le processus de différenciation des cellules souches hématopoïétiques en mégacaryocytes produisant les plaquettes sanguines. Des données de transcriptome indiquent la présence des récepteurs aux interleukines (IL) -10, -17A et -21 sur les mégacaryocytes. Mon travail montre que les IL-10 et -17 n’ont pas d’effet apparent sur la prolifération et la différenciation in vitro des mégacaryocytes à partir des progéniteurs hématopoïétiques humains CD34+. Cependant, l’IL-21 augmente la prolifération des progéniteurs mégacaryocytaires dérivés de ces cellules et sans modifier leur différenciation. Les mégacaryocytes de la moelle osseuse humaine expriment aussi le récepteur à l’IL-21, ce qui suggère un rôle direct de l’IL-21 sur la mégacaryocytopoïèse in vivo. De façon concordante, l’expression de l’IL-21 chez la souris stimule la mégacaryocytopoïèse ainsi que la production des plaquettes, mais augmente la clairance des plaquettes par les macrophages. Ces travaux suggèrent que durant les réponses immunitaires, l’expression de l’IL-21 par les lymphocytes T CD4+ activés module l’homéostasie des plaquettes sanguines.Megakaryocytopoiesis is the process by which hematopoietic stem cells give rise to megakaryocytes which in turn produce blood platelets. Transcriptional studies indicate the presence of interleukin (IL) -10, -17 and -21 receptors on megakaryocytes. This work shows that IL-10 and -17 have no apparent effect on in vitro proliferation and differentiation of megakaryocytes derived from human CD34+ hematopoietic progenitors. However, IL-21 increases the proliferation of megakaryocyte progenitors derived from these CD34+ cells without modifying their differentiation. Moreover, human bone marrow megakaryocytes express IL-21 receptor, suggesting a direct role of IL-21 on megakaryocytopoiesis in vivo. Concordantly, IL-21 expression in mice stimulates megakaryocytopoiesis and platelet production, but increases platelet clearance by macrophages. This work suggest that during immune responses, the expression of IL-21 by activated CD4+ T lymphocytes modulate blood platelet homeostasis

Dual role of IL-21 in megakaryopoiesis and platelet homeostasis: Modulation of platelet homeostasis by IL-21

International audienceGene profiling studies have indicated that in vitro differentiated human megakaryocytes express the receptor for IL-21 (IL-21R), an immunostimulatory cytokine associated with inflammatory disorders and currently under evaluation in cancer therapy. The aim of this study was to investigate whether IL-21 modulates megakaryopoiesis. We first checked the expression of IL-21 receptor on human BM and in vitro differentiated megakaryocytes. Then, we investigated the effect of IL-21 on the in vitro differentiation of human blood CD34+ progenitors into megakaryocytes. Finally, we analyzed the consequences of hydrodynamic transfection-mediated transient expression of IL-21, on megakaryopoiesis and thrombopoiesis in mice. The IL-21Rα chain was expressed in human BM megakaryocytes and was progressively induced during in vitro differentiation of human peripheral CD34+ progenitors, while the signal transducing γ chain was down-regulated. Consistently, the STAT3 phosphorylation induced by IL-21 diminished during the later stages of megakaryocytic differentiation. In vitro, IL-21 increased the number of CFU-MKs generated from CD34+ cells and the number of megakaryocytes differentiated from CD34+ progenitors in a JAK3- and STAT3-dependent manner. Forced expression of IL-21 in mice increased the density of bi-potent MK progenitors and BM megakaryocytes, and the platelet generation, but increased platelet clearance and consequently resulting in reduced blood platelet counts. Our work suggests that IL-21 regulates megakaryocyte development and platelet homeostasis. Thus IL-21 may link immune responses to physiological or pathological platelet-dependent processes

Single-cell genomics reveals the genetic and molecular bases for escape from mutational epistasis in myeloid neoplasms

Large-scale sequencing studies of hematologic malignancies have revealed notable epis-tasis among high-frequency mutations. One of the most striking examples of epistasis occurs for mutations in RNA splicing factors. These lesions are among the most common alterations in myeloid neoplasms and generally occur in a mutually exclusive manner, a finding attributed to their synthetic lethal interactions and/or convergent effects. Curiously, however, patients with multiple-concomitant splicing factor mutations have been observed, challenging our understanding of one of the most common examples of epistasis in hematologic malignancies. In this study, we performed bulk and single-cell analyses of patients with myeloid malignancy who were harboring double dagger 2 splicing factor mutations, to understand the frequency and basis for the coexistence of these mutations. Although mutations in splicing factors were strongly mutually exclusive across 4231 patients (q < .001), 0.85% harbored 2 concomitant bona fide splicing factor mutations, similar to 50% of which were present in the same individual cells. However, the distribution of mutations in patients with double mutations deviated from that in those with single mutations, with selection against the most common alleles, SF3B1K700E and SRSF2P95H/L/R, and selection for less common alleles, such as SF3B1 non-K700E mutations, rare amino acid substitutions at SRSF2P95, and combined U2AF1S34/Q157 mutations. SF3B1 and SRSF2 alleles enriched in those with double-mutations had reduced effects on RNA splicing and/or binding compared with the most common alleles. Moreover, dual U2AF1 mutations occurred in cis with preservation of the wild type allele. These data highlight allele-specific differences as critical in regulating the molecular effects of splicing factor mutations as well as their cooccurrences/exclusivities with one another

High caspase 3 and vulnerability to dual BCL2 family inhibition define ETO2::GLIS2 pediatric leukemia

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Impaired proteolysis of non-canonical RAS proteins drives clonal hematopoietic transformation

Recently, screens for mediators of resistance to FLT3 and ABL kinase inhibitors in leukemia resulted in the discovery of LZTR1 as an adapter of a Cullin-3 RING E3 ubiquitin ligase complex responsible for the degradation of RAS GTPases. In parallel, dysregulated LZTR1 expression via aberrant splicing and mutations was identified in clonal hematopoietic conditions. Here we identify that loss of LZTR1, or leukemia-associated mutants in the LZTR1 substrate and RAS GTPase RIT1 that escape degradation, drives hematopoietic stem cell (HSC) expansion and leukemia in vivo. Although RIT1 stabilization was sufficient to drive hematopoietic transformation, transformation mediated by LZTR1 loss required MRAS. Proteolysis targeting chimeras (PROTAC) against RAS or reduction of GTP-loaded RAS overcomes LZTR1 loss-mediated resistance to FLT3 inhibitors. These data reveal proteolysis of noncanonical RAS proteins as novel regulators of HSC self-renewal, define the function of RIT1 and LZTR1 mutations in leukemia, and identify means to overcome drug resistance due to LZTR1 downregulation.SignificanceHere we identify that impairing proteolysis of the noncanonical RAS GTPases RIT1 and MRAS via LZTR1 downregulation or leukemia-associated mutations stabilizing RIT1 enhances MAP kinase activation and drives leukemogenesis. Reducing the abundance of GTP-bound KRAS and NRAS overcomes the resistance to FLT3 kinase inhibitors associated with LZTR1 downregulation in leukemia. This article is highlighted in the In This Issue feature, p. 2221

Tumour spheres with inverted polarity drive the formation of peritoneal metastases in patients with hypermethylated colorectal carcinomas

International audienceMetastases account for 90% of cancer-related deaths; thus, it is vital to understand the biology of tumour dissemination. Here, we collected and monitored >50 patient specimens ex vivo to investigate the cell biology of colorectal cancer (CRC) metastatic spread to the peritoneum. This reveals an unpredicted mode of dissemination. Large clusters of cancer epithelial cells displaying a robust outward apical pole, which we termed tumour spheres with inverted polarity (TSIPs), were observed throughout the process of dissemination. TSIPs form and propagate through the collective apical budding of hypermethylated CRCs downstream of canonical and non-canonical transforming growth factor-β signalling. TSIPs maintain their apical-out topology and use actomyosin contractility to collectively invade three-dimensional extracellular matrices. TSIPs invade paired patient peritoneum explants, initiate metastases in mice xenograft models and correlate with adverse patient prognosis. Thus, despite their epithelial architecture and inverted topology TSIPs seem to drive the metastatic spread of hypermethylated CRCs