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MicroRNAs (miRNAs) are a class of endogenously encoded ~22 nucleotide, noncoding, single-stranded RNAs that contribute to development, body planning, stem cell differentiation, and tissue identity through posttranscriptional regulation and degradation of transcripts. Given their importance, it is predictable that dysregulation of miRNAs, which target a wide variety of transcripts, can result in malignant transformation. In this review, we explore the discovery of miRNAs, their mechanism of action, and the tools that aid in their discovery and study. Strikingly, many of the studies that have expanded our understanding of the contributions of miRNAs to normal physiology and in the development of diseases have come from studies in the hematopoietic system and hematologic malignancies, with some of the earliest identified functions for mammalian miRNAs coming from observations made in leukemias. So, with a special focus on the hematologic system, we will discuss how miRNAs contribute to differentiation of stem cells and how dysregulation of miRNAs contributes to the development of malignancy, by providing examples of specific miRNAs that function as oncogenes or tumor suppressors, as well as of defects in miRNA processing. Finally, we will discuss the promise of miRNA-based therapeutics and challenges for the future study of disease-causing miRNAs.

In the adult, the source of functionally diverse, mature blood cells are hematopoietic stem cells, a rare population of quiescent cells that reside in the bone marrow niche. Like stem cells in other tissues, hematopoietic stem cells are defined by their ability to self-renew, in order to maintain the stem cell population for the lifetime of the organism, and to differentiate, in order to give rise to the multiple lineages of the hematopoietic system. In recent years, increasing evidence has suggested a role for the accumulation of reactive oxygen species and DNA damage in the decision for hematopoietic stem cells to exit quiescence and to differentiate. In this review, we will examine recent work supporting the idea that detection of cell stressors, such as oxidative and genetic damage, is an important mediator of cell fate decisions in hematopoietic stem cells. We will explore the benefits of such a system in avoiding the development and progression of malignancies, and in avoiding tissue exhaustion and failure. Additionally, we will discuss new work that examines the accumulation of DNA damage and replication stress in aging hematopoietic stem cells and causes us to rethink ideas of genoprotection in the bone marrow niche.

The tumor suppressor promyelocytic leukemia (PML) was first identified as a component of PML–RARα fusion protein, one of the initiating cytogenetic abnormalities in acute promyelocytic leukemia. PML is now known to have diverse functions regulating the DNA-damage response, apoptosis, senescence, and angiogenesis. Recent investigations have identified PML as a regulator of metabolic pathways in stem cell compartments, including the hematopoietic system, and have provided researchers with new strategies for controlling stem cell maintenance and differentiation. Studies of PML in leukemia-initiating cells demonstrate that PML is also an essential component of their maintenance, which has drawn tremendous attention to PML from scientists in various stem cell fields. Here, we review research into PML and its associated pathways, including recent studies of PML as it relates to stem cell biology, as well as our finding that PML regulates fatty acid oxidation, which is essential to the maintenance of normal hematopoietic stem cells. We also discuss the therapeutic potential of controlling PML-associated pathways. In particular, we describe promising evidence for the use of arsenic trioxide in the treatment of chronic myeloid leukemia.

Precise control of microRNA expression contributes to development and the establishment of tissue identity, including in proper hematopoietic commitment and differentiation, whereas aberrant expression of various microRNAs has been implicated in malignant transformation. A small number of microRNAs are upregulated in megakaryocytes, among them is microRNA-22 (miR-22). Dysregulation of miR-22 leads to various hematologic malignancies and disorders, but its role in hematopoiesis is not yet well established. Here we show that upregulation of miR-22 is a critical step in megakaryocyte differentiation. Megakaryocytic differentiation in cell lines is promoted upon overexpression of miR-22, whereas differentiation is disrupted in CRISPR/Cas9-generated miR-22 knockout cell lines, confirming that miR-22 is an essential mediator of this process. RNA-sequencing reveals that miR-22 loss results in downregulation of megakaryocyte-associated genes. Mechanistically, we identify the repressive transcription factor, GFI1, as the direct target of miR-22, and upregulation of GFI1 in the absence of miR-22 inhibits megakaryocyte differentiation. Knocking down aberrant GFI1 expression restores megakaryocytic differentiation in miR-22 knockout cells. Furthermore, we have characterized hematopoiesis in miR-22 knockout animals and confirmed that megakaryocyte differentiation is similarly impaired in vivo and upon ex vivo megakaryocyte differentiation. Consistently, repression of Gfi1 is incomplete in the megakaryocyte lineage in miR-22 knockout mice and Gfi1 is aberrantly expressed upon forced megakaryocyte differentiation in explanted bone marrow from miR-22 knockout animals. This study identifies a positive role for miR-22 in hematopoiesis, specifically in promoting megakaryocyte differentiation through repression of GFI1, a target antagonistic to this process.

The ability of hematopoietic stem cells (HSCs) to self-renew and differentiate into progenitors is essential for homeostasis of the hematopoietic system. The longevity of HSCs makes them vulnerable to accumulating DNA damage, which may be leukemogenic or result in senescence and cell death. Additionally, the ability of HSCs to self-renew and differentiate allows DNA damage to spread throughout the hematologic system, leaving the organism vulnerable to disease. In this review we discuss cell fate decisions made in the face of DNA damage and other cellular stresses, and the role of reactive oxygen species in the long-term maintenance of HSCs and their DNA damage response.

The aryl hydrocarbon receptor (AhR) is a transcription factor involved in physiological processes, but also mediates most, if not all, toxic responses to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Activation of the AhR by TCDD leads to its dimerization with ARNT and transcriptional activation of several phase I and II metabolising enzymes. However, this classical signalling pathway so far failed to explain the pleiotrophic hazardous effects of TCDD such as developmental toxicity and tumour promotion. Thus, there is an urgent need to define genetic programmes orchestrated by AhR to unravel its role in physiology and toxicology. Treatment of rat liver oval cells with TCDD leads to a release from contact-inhibition. Loss of contact-inhibition is one characteristic hallmark in tumourigenesis. We have recently shown that TCDDexposure leads to an elevation of JunD protein levels and to transcriptional activation of Cyclin A in an AhRdependent, and probably ARNT-independent way. Ectopic expression of Cyclin A in confluent cultures overcomes G1-arrest indicating that increased Cyclin A levels are indeed sufficient to bypass contact-inhibition. Elevation of JunD precedes that of Cyclin A suggesting a role of JunD in Cyclin A induction. Indeed, down-regulating JunD by siRNA blocks TCDD-induced expression of Cyclin A. DNA affinity purification assays and reporter gene analysis indicate that JunD binds to an ATF/CRE consensus sequence in the rat Cyclin A promoter. Using in vitro DNA affinity purification assays, we also revealed binding of ATF2, but not Fra-or Fos-proteins, to the ATF/CRE consensus sequence. Down-regulating ATF2 by siRNA blocks TCDD-dependent Cyclin A induction indicating that ATF2 is the interaction partner of JunD mediating Cyclin A expression. In summary, we have discovered in rat liver oval cells a novel AhR-dependent and probably ARNTindependent signalling pathway involving JunD/ATF2 and Cyclin A, which mediates deregulation of contactinhibition by TCDD.

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