RESUMEN
MicroRNAs (miRNAs) play a crucial role in regulating gene expression. MicroRNA expression levels fluctuate, and point mutations and methylation occur in cancer cells; however, to date, there have been no reports of carcinogenic point mutations in miRNAs. MicroRNA 142 (miR-142) is frequently mutated in patients with follicular lymphoma, diffuse large B-cell lymphoma, chronic lymphocytic leukemia (CLL), and acute myeloid leukemia/myelodysplastic syndrome (AML/MDS). To understand the role of miR-142 mutation in blood cancers, the CRISPR-Cas9 system was utilized to successfully generate miR-142-55A>G mutant knock-in (Ki) mice, simulating the most frequent mutation in patients with miR-142 mutated AML/MDS. Bone marrow cells from miR-142 mutant heterozygous Ki mice were transplanted, and we found that the miR-142 mutant/wild-type cells were sufficient for the development of CD8+ T-cell leukemia in mice post-transplantation. RNA-sequencing analysis in hematopoietic stem/progenitor cells and CD8+ T-cells revealed that miR-142-Ki/+ cells had increased expression of the mTORC1 activator, a potential target of wild-type miR-142-3p. Notably, the expression of genes involved in apoptosis, differentiation, and the inhibition of the Akt-mTOR pathway was suppressed in miR-142-55A>G heterozygous cells, indicating that these genes are repressed by the mutant miR-142-3p. Thus, in addition to the loss of function due to the halving of wild-type miR-142-3p alleles, mutated miR-142-3p gained the function to suppress the expression of distinct target genes, sufficient to cause leukemogenesis in mice.
Asunto(s)
Leucemia Mieloide Aguda , MicroARNs , Síndromes Mielodisplásicos , Animales , Ratones , Carcinogénesis , Linfocitos T CD8-positivos/metabolismo , Mutación con Ganancia de Función , Leucemia Mieloide Aguda/genética , MicroARNs/genética , MicroARNs/metabolismo , Síndromes Mielodisplásicos/genéticaRESUMEN
Innate lymphoid cells are central to the regulation of immunity at mucosal barrier sites, with group 2 innate lymphoid cells (ILC2s) being particularly important in type 2 immunity. In this study, we demonstrate that microRNA(miR)-142 plays a critical, cell-intrinsic role in the homeostasis and function of ILC2s. Mice deficient for miR-142 expression demonstrate an ILC2 progenitor-biased development in the bone marrow, and along with peripheral ILC2s at mucosal sites, these cells display a greatly altered phenotype based on surface marker expression. ILC2 proliferative and effector functions are severely dysfunctional following Nippostrongylus brasiliensis infection, revealing a critical role for miR-142 isoforms in ILC2-mediated immune responses. Mechanistically, Socs1 and Gfi1 expression are regulated by miR-142 isoforms in ILC2s, impacting ILC2 phenotypes as well as the proliferative and effector capacity of these cells. The identification of these novel pathways opens potential new avenues to modulate ILC2-dependent immune functions.
Asunto(s)
Linfocitos/inmunología , MicroARNs/inmunología , Animales , Células HEK293 , Homeostasis , Humanos , Inmunidad Innata/inmunología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , MicroARNs/genéticaRESUMEN
The RUNX1 transcription factor is a critical regulator of normal haematopoiesis and its functional disruption by point mutations, deletions or translocations is a major causative factor leading to leukaemia. In the majority of cases, genetic changes in RUNX1 are linked to loss of function classifying it broadly as a tumour suppressor. Despite this, several recent studies have reported the need for a certain level of active RUNX1 for the maintenance and propagation of acute myeloid leukaemia and acute lymphoblastic leukaemia cells, suggesting an oncosupportive role of RUNX1. Furthermore, in solid cancers, RUNX1 is overexpressed compared with normal tissue, and RUNX factors have recently been discovered to promote growth of skin, oral, breast and ovarian tumour cells, amongst others. RUNX factors have key roles in stem cell fate regulation during homeostasis and regeneration of many tissues. Cancer cells appear to have corrupted these stem cell-associated functions of RUNX factors to promote oncogenesis. Here, we discuss current knowledge on the role of RUNX genes in stem cells and as oncosupportive factors in haematological malignancies and epithelial cancers.
Asunto(s)
Subunidad alfa 2 del Factor de Unión al Sitio Principal/metabolismo , Modelos Moleculares , Proteínas de Neoplasias/metabolismo , Neoplasias/metabolismo , Células Madre/metabolismo , Animales , Carcinogénesis , Diferenciación Celular , Subunidad alfa 2 del Factor de Unión al Sitio Principal/química , Subunidad alfa 2 del Factor de Unión al Sitio Principal/genética , Regulación del Desarrollo de la Expresión Génica , Regulación Neoplásica de la Expresión Génica , Humanos , Mutación , Proteínas de Neoplasias/química , Proteínas de Neoplasias/genética , Neoplasias/patología , Células Madre/citología , Células Madre/patologíaRESUMEN
A lentiviral vector (LV) pseudotype derived from the fusion (F) and hemagglutinin-neuraminidase (HN) glycoproteins of a murine respirovirus (Sendai virus) facilitates efficient targeting of murine lung in vivo. Since targeting of the human lung will depend upon the availability and distribution of receptors used by F/HN, we investigated transduction of primary human airway cells differentiated at the air-liquid interface (ALI). We observed targeting of human basal, ciliated, goblet, and club cells, and using a combination of sialidase enzymes and lectins, we showed that transduction is dependent on the availability of sialylated glycans, including α2,3 sialylated N-acetyllactosamine (LacNAc). Transduction via F/HN was 300-fold more efficient than another hemagglutinin-based LV pseudotype derived from influenza fowl plague virus (HA Rostock), despite similar efficiency reported in murine airways in vivo. Using specific glycans to inhibit hemagglutination, we showed this could be due to a greater affinity of F/HN for α2,3 sialylated LacNAc. Overall, these results highlight the importance of identifying the receptors used in animal and cell-culture models to predict performance in the human airways. Given the reported prevalence of α2,3 sialylated LacNAc on human pulmonary cells, these results support the suitability of the F/HN pseudotype for human lung gene therapy applications.
RESUMEN
The ETV6-RUNX1 onco-fusion arises in utero, initiating a clinically silent pre-leukemic state associated with the development of pediatric B-acute lymphoblastic leukemia (B-ALL). We characterize the ETV6-RUNX1 regulome by integrating chromatin immunoprecipitation- and RNA-sequencing and show that ETV6-RUNX1 functions primarily through competition for RUNX1 binding sites and transcriptional repression. In pre-leukemia, this results in ETV6-RUNX1 antagonization of cell cycle regulation by RUNX1 as evidenced by mass cytometry analysis of B-lineage cells derived from ETV6-RUNX1 knock-in human pluripotent stem cells. In frank leukemia, knockdown of RUNX1 or its co-factor CBFß results in cell death suggesting sustained requirement for RUNX1 activity which is recapitulated by chemical perturbation using an allosteric CBFß-inhibitor. Strikingly, we show that RUNX1 addiction extends to other genetic subtypes of pediatric B-ALL and also adult disease. Importantly, inhibition of RUNX1 activity spares normal hematopoiesis. Our results suggest that chemical intervention in the RUNX1 program may provide a therapeutic opportunity in ALL.
Asunto(s)
Subunidad alfa 2 del Factor de Unión al Sitio Principal , Leucemia-Linfoma Linfoblástico de Células Precursoras , Adulto , Niño , Humanos , Subunidad alfa 2 del Factor de Unión al Sitio Principal/genética , Factores de Unión al Sitio Principal , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras/terapia , Linfocitos B , Fusión GénicaRESUMEN
Defective silencing of retrotransposable elements has been linked to inflammageing, cancer and autoimmune diseases. However, the underlying mechanisms are only partially understood. Here we implicate the histone H3.3 chaperone Daxx, a retrotransposable element repressor inactivated in myeloid leukaemia and other neoplasms, in protection from inflammatory disease. Loss of Daxx alters the chromatin landscape, H3.3 distribution and histone marks of haematopoietic progenitors, leading to engagement of a Pu.1-dependent transcriptional programme for myelopoiesis at the expense of B-cell differentiation. This causes neutrophilia and inflammation, predisposing mice to develop an autoinflammatory skin disease. While these molecular and phenotypic perturbations are in part reverted in animals lacking both Pu.1 and Daxx, haematopoietic progenitors in these mice show unique chromatin and transcriptome alterations, suggesting an interaction between these two pathways. Overall, our findings implicate retrotransposable element silencing in haematopoiesis and suggest a cross-talk between the H3.3 loading machinery and the pioneer transcription factor Pu.1.
Asunto(s)
Cromatina/patología , Proteínas Co-Represoras/genética , Trastornos Leucocíticos/congénito , Chaperonas Moleculares/genética , Mielopoyesis/genética , Proteínas Proto-Oncogénicas/metabolismo , Transactivadores/metabolismo , Animales , Enfermedades Autoinmunes/genética , Enfermedades Autoinmunes/patología , Linfocitos B/citología , Línea Celular , Cromatina/genética , Células Madre Hematopoyéticas/citología , Histonas/metabolismo , Humanos , Inflamación/patología , Trastornos Leucocíticos/patología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Retroelementos/genética , Enfermedades de la Piel/genética , Enfermedades de la Piel/inmunología , Enfermedades de la Piel/patologíaRESUMEN
MicroRNAs (miRNAs) were first discovered in genetic screens for regulators of developmental timing in the stem-cell-like seam cell lineage in Caenorhabditis elegans. As members of the heterochronic pathway, the lin-4 and let-7 miRNAs are required in the seam cells for the correct progression of stage-specific events and to ensure that cell cycle exit and terminal differentiation occur at the correct time. Other heterochronic genes such as lin-28 and lin-41 are direct targets of the lin-4 and let-7 miRNAs. Recent findings on the functions of the let-7 and lin-4/mir-125 miRNA families and lin-28 and lin-41 orthologs from a variety of organisms suggest that core elements of the heterochronic pathway are retained in mammalian stem cells and development. In particular, these genes appear to form bistable switches via double-negative feedback loops in both nematode and mammalian stem cell development, the functional relevance of which is finally becoming clear. let-7 inhibits stem cell self-renewal in both normal and cancer stem cells of the breast and acts as a tumor suppressor in lung and breast cancer. let-7 also promotes terminal differentiation at the larval to adult transition in both nematode stem cells and fly wing imaginal discs and inhibits proliferation of human lung and liver cancer cells. Conversely, LIN-28 is a highly specific embryonic stem cell marker and is one of four "stemness" factors used to reprogram adult fibroblasts into induced pluripotent stem cells; furthermore, lin-28 is oncogenic in hepatocellular carcinomas. Therefore, a core module of heterochronic genes--lin-28, lin-41, let-7, and lin-4/mir-125-acts as an ancient regulatory switch for differentiation in stem cells (and in some cancers), illustrating that nematode seam cells mirror miRNA regulatory networks in mammalian stem cells during both normal development and cancer.
Asunto(s)
Caenorhabditis elegans/genética , MicroARNs/genética , Células Madre/metabolismo , Animales , Caenorhabditis elegans/citología , Caenorhabditis elegans/crecimiento & desarrollo , Regulación del Desarrollo de la Expresión Génica , MicroARNs/metabolismo , Modelos Biológicos , Neoplasias/genética , Neoplasias/patología , Células Madre/citología , Factores de TiempoRESUMEN
Runx family transcription factors have risen to prominence over the last few years because of the increasing evidence implicating them as key regulators of the choice between cell proliferation and differentiation during development and carcinogenesis. Runx factors have been found to be involved in diverse developmental processes, ranging from hematopoiesis to neurogenesis, and are increasingly being linked with various human cancers. In this review, we examine the case for Runx factors as key regulators of cell proliferation in various developmental situations, a role that predisposes Runx mutations as causative agents in oncogenesis. We discuss the evidence that Runx factors regulate, and are regulated by, core components of the cell cycle machinery, and focus our attention on the solo Runx gene, rnt-1, in Caenorhabditis elegans, an organism that we feel has much to offer the Runx field.
Asunto(s)
Biología , Proteínas de Caenorhabditis elegans/fisiología , Caenorhabditis elegans/fisiología , Subunidades alfa del Factor de Unión al Sitio Principal/fisiología , Animales , Caenorhabditis elegans/citología , Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/genética , Ciclo Celular , Diferenciación Celular , Linaje de la Célula , Proliferación Celular , Subunidades alfa del Factor de Unión al Sitio Principal/genética , Mutación , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/fisiología , Neoplasias/genética , Proteínas Proto-Oncogénicas/fisiología , Células Madre/citología , Células Madre/fisiología , Factores de Transcripción/genética , Factores de Transcripción/fisiologíaRESUMEN
Regulation of lineage commitment in multipotential cells is key to maintaining a balanced hematopoietic output throughout life while retaining the capacity to respond to stress and infection. Cell fate decisions are made by individual stem cells, but population-level analysis obscures the mechanics of cell fate choice by averaging the molecular and functional heterogeneity that exists even in the most highly purified stem cell populations. Therefore, single cell analysis of both molecular and cellular phenotypes is crucial to delineate and interrogate the process of lineage commitment. We review recent single cell expression profiling, imaging, and clonal tracking studies that have provided new insights into commitment, focusing on the hematopoietic system, and suggest how new technologies may illuminate our understanding of lineage commitment in the near future.
Asunto(s)
Diferenciación Celular/fisiología , Linaje de la Célula/fisiología , Redes Reguladoras de Genes/fisiología , Análisis de la Célula Individual/métodos , Animales , Humanos , Análisis de la Célula Individual/tendenciasRESUMEN
Hematopoietic stem cells (HSCs) emerge during embryogenesis from hemogenic endothelium, but it remains unclear how the HSC lineage is initially established from mesoderm during ontogeny. In Xenopus, the definitive hemangioblast precursors of the HSC lineage have been identified in dorsal lateral plate (DLP) mesoderm, and a transcriptional gene regulatory network (GRN) controlling hemangioblast programming has been elucidated. Herein, we identify an essential role for microRNAs (miRNAs) in establishing the mesodermal lineage leading to both HSC emergence and vasculogenesis and determine that a single miRNA, miR-142-3p, is primarily responsible for initiation of definitive hemangioblast specification. miR-142-3p forms a double-negative gate unlocking entry into the hemangioblast program, in part by inhibiting TGFß signaling. Our results table miR-142-3p as a master regulator of HSC lineage specification, sitting at the apex of the hierarchy programming the adult hemangioblast, thus illustrating that miRNAs can act as instructive determinants of cell fate during development.
Asunto(s)
Regulación del Desarrollo de la Expresión Génica , Hemangioblastos/fisiología , Células Madre Hematopoyéticas/fisiología , MicroARNs/fisiología , Neovascularización Fisiológica/genética , Animales , Diferenciación Celular , Linaje de la Célula/fisiología , Desarrollo Embrionario/fisiología , Mesodermo/citología , Ratones , Ratones Endogámicos C57BL , MicroARNs/genética , Morfolinos , Proteínas/genética , Proteínas de Unión al ARN , Xenopus laevisRESUMEN
In this report, we investigate the C. elegans CBFbeta homologue, BRO-1. bro-1 mutants have a similar male-specific sensory ray loss phenotype to rnt-1 (the C. elegans homologue of the mammalian CBFbeta-interacting Runx factors), caused by failed cell divisions in the seam lineages. Our studies indicate that BRO-1 and RNT-1 form a cell proliferation-promoting complex, and that BRO-1 increases both the affinity and specificity of RNT-1-DNA interactions. Overexpression of bro-1, like rnt-1, leads to an expansion of seam cell number and co-overexpression of bro-1 and rnt-1 results in massive seam cell hyperplasia. Finally, we find that BRO-1 appears to act independently of RNT-1 in certain situations. These studies provide new insights into the function and regulation of this important cancer-associated DNA-binding complex in stem cells and support the view that Runx/CBFbeta factors have oncogenic potential.
Asunto(s)
Proteínas de Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/citología , Caenorhabditis elegans/metabolismo , Proteínas Represoras/metabolismo , Células Madre/citología , Células Madre/metabolismo , Factores de Transcripción/metabolismo , Secuencia de Aminoácidos , Animales , Caenorhabditis elegans/genética , Caenorhabditis elegans/crecimiento & desarrollo , Proteínas de Caenorhabditis elegans/genética , Linaje de la Célula , Proliferación Celular , ADN/metabolismo , Femenino , Regulación del Desarrollo de la Expresión Génica , Humanos , Masculino , Datos de Secuencia Molecular , Mutación/genética , Fenotipo , Unión Proteica , Proteínas Represoras/genética , Alineación de Secuencia , Homología de Secuencia de Aminoácido , Factores de Transcripción/genética , Regulación hacia ArribaRESUMEN
In this report, we demonstrate that C. elegans mab-2 mutants have defects in the development of a male-specific sense organ because of a failure in the proliferation of the stem cell-like lateral hypodermal (seam) cells. We show, by positional cloning, that mab-2 encodes RNT-1, the only C. elegans member of the Runx family of transcriptional regulators, which are postulated to act both as oncogenes and tumour suppressors in mammalian cells. Importantly, we find that rnt-1 is a rate-limiting regulator of seam cell proliferation in C. elegans, as overexpression of rnt-1 at particular developmental stages is capable of driving extra cell divisions, leading to seam cell hyperplasia. Loss of rnt-1 is correlated with upregulation of cki-1, a CDK inhibitor. Deregulated expression of Runx genes in humans is associated with various cancers, particularly leukaemias, suggesting a conserved role for Runx genes in controlling cell proliferation during development, especially in stem cell lineages. C. elegans is therefore an important model system for studying the biology, and oncogenic potential, of Runx genes.
Asunto(s)
Proteínas de Caenorhabditis elegans/fisiología , Caenorhabditis elegans/fisiología , Proliferación Celular , Células Madre/citología , Factores de Transcripción/fisiología , Animales , Caenorhabditis elegans/citología , Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/genética , Linaje de la Célula/fisiología , Subunidades alfa del Factor de Unión al Sitio Principal/genética , Femenino , Masculino , Interferencia de ARN , Células Madre/fisiología , Factores de Transcripción/genéticaRESUMEN
A recent report by Blumenthal et al. provides convincing evidence that at least 15% of Caenorhabditis elegans genes are co-transcribed within over a thousand operons. Polycistronic transcription of gene clusters is very rare in eukaryotes. The widespread occurrence of operons in C. elegans thus raises some interesting questions about the origin and function of these multigenic transcriptional units.