Stop the dicing in hematopoiesis: What have we learned?

MicroRNAs (miRNAs) belong to an abundant class of highly conserved small (22nt) non-coding RNAs. MiRNA profiling studies indicate that their expression is highly cell type-dependent. DICER1 is an essential RNase III endoribonuclease for miRNA processing. Hematopoietic cell type- and developmental stage-specific Dicer1 deletion models show that miRNAs are essential regulators of cellular survival, differentiation and function. For instance, miRNA deficiency in hematopoietic stem cells and progenitors of different origins results in decreased cell survival, dramatic developmental aberrations or dysfunctions in mice. We recently found that homozygous Dicer1 deletion in myeloid-committed progenitors results in an aberrant expression of stem cell genes and induces a regained self-renewal capacity. Moreover, Dicer1 deletion causes a block in macrophage development and myeloid dysplasia, a cellular condition that may be considered as a preleukemic state. However, Dicer1-null cells do not develop leukemia in mice, indicating that depletion of miRNAs is not enough for tumorigenesis. Surprisingly, we found that heterozygous Dicer1 deletion in myeloid-committed progenitors, but not Dicer1 knockout, collaborates with p53 deletion in leukemic progression and results in various types of leukemia. Our data indicate that Dicer1 is a haploinsufficient tumorsuppressor in hematopoietic neoplasms, which is consistent with the observed downregulation of miRNA expression in human leukemia samples. Here, we review the various hematopoietic specific Dicer1 deletion mouse models and the phenotypes observed within the different hematopoietic lineages and cell developmental stages. Finally, we discuss the role for DICER1 in mouse and human malignant hematopoiesis

The interplay between critical transcription factors and microRNAs in the control of normal and malignant myelopoiesis

Myelopoiesis is a complex process driven by essential transcription factors, including C/EBPα, PU.1, RUNX1, KLF4 and IRF8. Together, these factors are critical for the control of myeloid progenitor cell expansion and lineage determination in the development of granulocytes and monocytes/macrophages. MicroRNAs (miRNAs) are expressed in a cell type and lineage specific manner. There is increasing evidence that miRNAs fine-tune the expression of hematopoietic lineage-specific transcription factors and drive the lineage decisions of hematopoietic progenitor cells. In this review, we discuss recently discovered self-activating and feed-back mechanisms in which transcription factors and miRNAs interact during myeloid cell development. Furthermore, we delineate how some of these mechanisms are affected in acute myeloid leukemia (AML) and how disrupted transcription factor-miRNA interplays contribute to leukemogenesis

A genome-wide scan for microrna-related genetic variants associated with primary open-angle glaucoma

PURPOSE: To identify microRNAs (miRNAs) involved in primary open-angle glaucoma (POAG), using genetic data. MiRNAs are small noncoding RNAs that posttranscriptionally regulate gene expression. Genetic variants in miRNAs or miRNA-binding sites within gene 3’-untranslated regions (3’UTRs) are expected to affect miRNA function and con

Identification of osteolineage cell-derived extracellular vesicle cargo implicated in hematopoietic support

Osteolineage cell-derived extracellular vesicles (EVs) play a regulatory role in hematopoiesis and have been shown to promote the ex vivo expansion of human hematopoietic stem and progenitor cells (HSPCs). Here, we demonstrate that EVs from different human osteolineage sources do not have the same HSPC expansion promoting potential. Comparison of stimulatory and non-stimulatory osteolineage EVs by next-generation sequencing and mass spectrometry analyses revealed distinct microRNA and protein signatures identifying EV-derived candidate regulators of ex vivo HSPC expansion. Accordingly, the treatment of umbilical cord blood-derived CD34+ HSPCs with stimulatory EVs-altered HSPC transcriptome, including genes with known roles in cell proliferation. An integrative bioinformatics approach, which connects the HSPC gene expression data with the candidate cargo in stimulatory EVs, delineated the potentially targeted biological functions and pathways during hematopoietic cell expansion and development. In conclusion, our study giv

Rapid in vitro generation of bona fide exhausted CD8+ T cells is accompanied by Tcf7 promotor methylation

Exhaustion is a dysfunctional state of cytotoxic CD8+ T cells (CTL) observed in chronic infection and cancer. Current in vivo models of CTL exhaustion using chronic viral infections or cancer yield very few exhausted CTL, limiting the analysis that can be done on these cells. Establishing an in vitro system that rapidly induces CTL exhaustion would therefore greatly facilitate the study of this phenotype, identify the truly exhaustion-associated changes and allow the testing of novel approaches to reverse or prevent exhaustion. Here we show that repeat stimulation of purified TCR transgenic OT-I CTL with their specific peptide induces all the functional (reduced cytokine production and polyfunctionality, decreased in vivo expansion capacity) and phenotypic (increased inhibitory receptors expression and transcription factor changes) characteristics of exhaustion. Importantly, in vitro exhausted cells shared the transcriptomic characteristics of the gold standard of exhaustion, CTL from LCMV cl13 infections. Gene expression of both in vitro and in vivo exhausted CTL was distinct from T cells anergy. Using this system, we show that Tcf7 promoter DNA methylation contributes to TCF1 downregulation in exhausted CTL. Thus this novel in vitro system can be used to identify genes and signaling pathways involved in exhaustion and will facilitate the screening of reagents that prevent/reverse CTL exhaustion

Identification, function and clinical relevance of myeloid leukemia genes

Acute myeloïde leukemie (AML) wordt geclassificeerd in verschillende risico categorieën op basis van cytogenetische en moleculaire afwijkingen. De chromosomale translocaties t(8;21), t(15;17) en inv(16) bijvoorbeeld, resulteren in fusie-eiwitten met afwijkende transcriptioneel regulerende activiteiten

The gene encoding the transcriptional regulator Yin Yang 1 (YY1) is a myeloid transforming gene interfering with neutrophilic differentiation

The genetic defects underlying the pathogenesis of acute myeloid leukemia (AML) are still largely unknown. Retroviral insertion mutagenesis in mice has become a powerful tool to identify candidate genes involved in the development of leukemia and lymphoma. We have used this strategy with the 1.4 strain of Graffi murine leukemia virus (MuLV), which predominantly causes myeloid leukemias. Here, we report that Graffi-1.4-induced AML frequently harbors virus integrations in the gene encoding the transcription factor Yin Yang 1 (YY1). These integrations occurred in both orientations, and all were located in the 5' promoter region of the gene, 0.5 to 1.5 kb upstream of the major transcriptional start site. Luciferase reporter assays showed

Dicer1 deletion in myeloid-committed progenitors causes neutrophil dysplasia and blocks macrophage/dendritic cell development in mice

MicroRNAs (miRNAs) have the potential to regulate cellular differentiation programs; however, miRNA deficiency in primary hematopoietic stem cells (HSCs) results in HSC depletion in mice, leaving the question of whether miRNAs play a role in early-lineage decisions unanswered. To address this issue, we deleted Dicer1, which encodes an essential RNase III enzyme for miRNA biogenesis, in murine CCAAT/enhancer-binding protein α (C/EBPA)-positive myeloid-committed progenitors in vivo. In contrast to the results in HSCs, we found that miRNA depletion affected neither the number of myeloid progenitors nor the percentage of C/EBPA-positive progenitor cells. Analysis of gene-expression profiles from wild-type and Dicer1-deficient granulocyte-macrophage progenitors (GMPs) revealed that 20 miRNA families were active in GMPs. Of the derepressed miRNA targets in Dicer1-null GMPs, 27% are normally exclusively expressed in HSCs or are specific for multipotent progenitors and erythropoiesis, indicating an altered geneexpression landscape. Dicer1-deficient GMPs were defective in myeloid development in vitro and exhibited an increased replating capacity, indicating the regained self-renewal potential of these cells. In mice, Dicer1 deletion blocked monocytic differentiation, depleted macrophages, and caused myeloid dysplasia with morphologic features of Pelger-Huë t anomaly. These results provide evidence for a miRNA-controlled switch for a cellular program of self-renewal and expansion toward myeloid differentiation in GMPs

A Genetic variant in the seed region of miR-4513 shows pleiotropic effects on lipid and glucose homeostasis, blood pressure, and coronary artery disease

MicroRNAs (miRNA) play a crucial role in the regulation of diverse biological processes by post-transcriptional modulation of gene expression. Genetic polymorphisms in miRNA-related genes can potentially contribute to a wide range of phenotypes. The effect of such variants on cardiometabolic diseases has not yet been defined. We systematically investigated the association of genetic variants in the seed region of miRNAs with cardiometabolic phenotypes, using the thus far largest genome-wide association studies on 17 cardiometabolic traits/diseases. We found that rs2168518:G>A, a seed region variant of miR-4513, associates with fasting glucose, low-density lipoprotein-cholesterol, total cholesterol, systolic and diastolic blood pressure, and risk of coronary artery disease. We experimentally showed that miR-4513 expression is significantly reduced in the presence of the rs2168518 mutant allele. We sought to identify miR-451

An autonomous CEBPA enhancer specific for myeloid-lineage priming and neutrophilic differentiation

Neutrophilic differentiation is dependent on CCAAT enhancer-binding protein α (C/EBPα), a transcription factor expressed in multiple organs including the bone marrow. Using functional genomic technologies in combination with clustered regularly-interspaced short palindromic repeat (CRISPR)/CRISPR-associated protein 9 genome editing and in vivo mouse modeling, we show that CEBPA is located in a 170-kb topological-associated domain that contains 14 potential enhancers. Of these, 1 enhancer located +42 kb from CEBPA is active and engages with the CEBPA promoter in myeloid cells only. Germ line deletion of the homologous enhancer in mice in vivo reduces Cebpa levels exclusively in hematopoietic stem cells (HSCs) and myeloid-primed progenitor cells leading to severe defects in the granulocytic lineage, without affecting any other Cebpa-expressing organ studied. The enhancer-deleted progenitor cells lose their myeloid transcription program and are blocked in differentiation. Deletion of the enhancer also causes loss of HSC maintenance. We conclude that a single +42-kb enhancer is essential for CEBPA expression in myeloid cells only