Induction of microRNAs, mir-155, mir-222, mir-424 and mir-503, promotes monocytic differentiation through combinatorial regulation

Acute myeloid leukemia (AML) involves a block in terminal differentiation of the myeloid lineage and uncontrolled proliferation of a progenitor state. Using phorbol myristate acetate (PMA), it is possible to overcome this block in THP-1 cells (an M5-AML containing the MLL-MLLT3 fusion), resulting in differentiation to an adherent monocytic phenotype. As part of FANTOM4, we used microarrays to identify 23 microRNAs that are regulated by PMA. We identify four PMA-induced micro- RNAs (mir-155, mir-222, mir-424 and mir-503) that when overexpressed cause cell-cycle arrest and partial differentiation and when used in combination induce additional changes not seen by any individual microRNA. We further characterize these prodifferentiative microRNAs and show that mir-155 and mir-222 induce G2 arrest and apoptosis, respectively. We find mir-424 and mir-503 are derived from a polycistronic precursor mir-424-503 that is under repression by the MLL-MLLT3 leukemogenic fusion. Both of these microRNAs directly target cell-cycle regulators and induce G1 cell-cycle arrest when overexpressed in THP-1. We also find that the pro-differentiative mir-424 and mir-503 downregulate the anti-differentiative mir-9 by targeting a site in its primary transcript. Our study highlights the combinatorial effects of multiple microRNAs within cellular systems.Comment: 45 pages 5 figure

Induction in myeloid leukemic cells of genes that are expressed in different normal tissues

Using DNA microarray and cluster analysis of expressed genes in a cloned line (M1-t-p53) of myeloid leukemic cells, we have analyzed the expression of genes that are preferentially expressed in different normal tissues. Clustering of 547 highly expressed genes in these leukemic cells showed 38 genes preferentially expressed in normal hematopoietic tissues and 122 other genes preferentially expressed in different normal non-hematopoietic tissues including neuronal tissues, muscle, liver and testis. We have also analyzed the genes whose expression in the leukemic cells changed after activation of wild-type p53 and treatment with the cytokine interleukin 6 (IL-6) or the calcium mobilizer thapsigargin (TG). Out of 620 such genes in the leukemic cells that were differentially expressed in normal tissues, clustering showed 80 genes that were preferentially expressed in hematopoietic tissues and 132 genes in different normal non-hematopietic tissues that also included neuronal tissues, muscle, liver and testis. Activation of p53 and treatment with IL-6 or TG induced different changes in the genes preferentially expressed in these normal tissues. These myeloid leukemic cells thus express genes that are expressed in normal non-hematopoietic tissues, and various treatments can reprogram these cells to induce other such non-hematopoietic genes. The results indicate that these leukemic cells share with normal hematopoietic stem cells the plasticity of differentiation to different cell types. It is suggested that this reprogramming to induce in malignant cells genes that are expressed in different normal tissues may be of clinical value in therapy

QoS Analysis in Heterogeneous Choreography Interactions

International audienceWith an increasing number of services and devices interacting in a decentralized manner, choreographies are an active area of investigation. The heterogeneous nature of interacting systems leads to choreographies that may not only include conventional services, but also sensor-actuator networks, databases and service feeds. Their middleware behavior within choreographies is captured through abstract interaction paradigms such as client-service, publish-subscribe and tuple space. In this paper, we study these heterogeneous interaction paradigms, connected through an eXtensible Service Bus proposed in the CHOReOS project. As the functioning of such choreographies is dependent on the Quality of Service (QoS) performance of participating entities, an intricate analysis of interaction paradigms and their effect on QoS metrics is needed. We study the composition of QoS metrics in heterogeneous choreographies, and the subsequent tradeoffs. This produces interesting insights such as selection of a particular system and its middleware during design time or end-to-end QoS expectation/guarantees during runtime. Non-parametric hypothesis tests are applied to systems, where QoS dependent services may be replaced at runtime to prevent deterioration in performance

Detection of microRNA Expression in Human Peripheral Blood Microvesicles

MicroRNAs (miRNA) are small non-coding RNAs that regulate translation of mRNA and protein. Loss or enhanced expression of miRNAs is associated with several diseases, including cancer. However, the identification of circulating miRNA in healthy donors is not well characterized. Microvesicles, also known as exosomes or microparticles, circulate in the peripheral blood and can stimulate cellular signaling. In this study, we hypothesized that under normal healthy conditions, microvesicles contain miRNAs, contributing to biological homeostasis.Microvesicles were isolated from the plasma of normal healthy individuals. RNA was isolated from both the microvesicles and matched mononuclear cells and profiled for 420 known mature miRNAs by real-time PCR. Hierarchical clustering of the data sets indicated significant differences in miRNA expression between peripheral blood mononuclear cells (PBMC) and plasma microvesicles. We observed 71 miRNAs co-expressed between microvesicles and PBMC. Notably, we found 33 and 4 significantly differentially expressed miRNAs in the plasma microvesicles and mononuclear cells, respectively. Prediction of the gene targets and associated biological pathways regulated by the detected miRNAs was performed. The majority of the miRNAs expressed in the microvesicles from the blood were predicted to regulate cellular differentiation of blood cells and metabolic pathways. Interestingly, a select few miRNAs were also predicted to be important modulators of immune function.This study is the first to identify and define miRNA expression in circulating plasma microvesicles of normal subjects. The data generated from this study provides a basis for future studies to determine the predictive role of peripheral blood miRNA signatures in human disease and will enable the definition of the biological processes regulated by these miRNA

Distinctive Patterns of MicroRNA Expression Associated with Karyotype in Acute Myeloid Leukaemia

Acute myeloid leukaemia (AML) is the most common acute leukaemia in adults; however, the genetic aetiology of the disease is not yet fully understood. A quantitative expression profile analysis of 157 mature miRNAs was performed on 100 AML patients representing the spectrum of known karyotypes common in AML. The principle observation reported here is that AMLs bearing a t(15;17) translocation had a distinctive signature throughout the whole set of genes, including the up regulation of a subset of miRNAs located in the human 14q32 imprinted domain. The set included miR-127, miR-154, miR-154*, miR-299, miR-323, miR-368, and miR-370. Furthermore, specific subsets of miRNAs were identified that provided molecular signatures characteristic of the major translocation-mediated gene fusion events in AML. Analysis of variance showed the significant deregulation of 33 miRNAs across the leukaemic set with respect to bone marrow from healthy donors. Fluorescent in situ hybridisation analysis using miRNA-specific locked nucleic acid (LNA) probes on cryopreserved patient cells confirmed the results obtained by real-time PCR. This study, conducted on about a fifth of the miRNAs currently reported in the Sanger database (microrna.sanger.ac.uk), demonstrates the potential for using miRNA expression to sub-classify cancer and suggests a role in the aetiology of leukaemia

Integration of Expressed Sequence Tag Data Flanking Predicted RNA Secondary Structures Facilitates Novel Non-Coding RNA Discovery

Many computational methods have been used to predict novel non-coding RNAs (ncRNAs), but none, to our knowledge, have explicitly investigated the impact of integrating existing cDNA-based Expressed Sequence Tag (EST) data that flank structural RNA predictions. To determine whether flanking EST data can assist in microRNA (miRNA) prediction, we identified genomic sites encoding putative miRNAs by combining functional RNA predictions with flanking ESTs data in a model consistent with miRNAs undergoing cleavage during maturation. In both human and mouse genomes, we observed that the inclusion of flanking ESTs adjacent to and not overlapping predicted miRNAs significantly improved the performance of various methods of miRNA prediction, including direct high-throughput sequencing of small RNA libraries. We analyzed the expression of hundreds of miRNAs predicted to be expressed during myogenic differentiation using a customized microarray and identified several known and predicted myogenic miRNA hairpins. Our results indicate that integrating ESTs flanking structural RNA predictions improves the quality of cleaved miRNA predictions and suggest that this strategy can be used to predict other non-coding RNAs undergoing cleavage during maturation

Epigenetic abnormalities in myeloproliferative neoplasms: a target for novel therapeutic strategies

The myeloproliferative neoplasms (MPNs) are a group of clonal hematological malignancies characterized by a hypercellular bone marrow and a tendency to develop thrombotic complications and to evolve to myelofibrosis and acute leukemia. Unlike chronic myelogenous leukemia, where a single disease-initiating genetic event has been identified, a more complicated series of genetic mutations appear to be responsible for the BCR-ABL1-negative MPNs which include polycythemia vera, essential thrombocythemia, and primary myelofibrosis. Recent studies have revealed a number of epigenetic alterations that also likely contribute to disease pathogenesis and determine clinical outcome. Increasing evidence indicates that alterations in DNA methylation, histone modification, and microRNA expression patterns can collectively influence gene expression and potentially contribute to MPN pathogenesis. Examples include mutations in genes encoding proteins that modify chromatin structure (EZH2, ASXL1, IDH1/2, JAK2V617F, and IKZF1) as well as epigenetic modification of genes critical for cell proliferation and survival (suppressors of cytokine signaling, polycythemia rubra vera-1, CXC chemokine receptor 4, and histone deacetylase (HDAC)). These epigenetic lesions serve as novel targets for experimental therapeutic interventions. Clinical trials are currently underway evaluating HDAC inhibitors and DNA methyltransferase inhibitors for the treatment of patients with MPNs

Transcriptional profiling identifies differential expression of long non-coding RNAs in Jo-1 associated and inclusion body myositis.

Myositis is characterised by muscle inflammation and weakness. Although generally thought to be driven by a systemic autoimmune response, increasing evidence suggests that intrinsic changes in the muscle might also contribute to the pathogenesis. Long non-coding RNAs (lncRNAs) are a family of novel genes that regulate gene transcription and translation. To determine the potential role of lncRNAs, we employed next generation sequencing to examine the transcriptome in muscle biopsies obtained from two histologically distinct patient populations, inclusion body myositis (IBM) and anti-Jo-1-associated myositis (Jo-1). 1287 mRNAs and 1068 mRNAs were differentially expressed in the muscle from Jo-1 and IBM patients, respectively. Pathway analysis showed the top canonical pathway in both Jo-1 and IBM was oxidative phosphorylation and mitochondrial dysfunction. We identified 731 known and 325 novel lncRNAs in the muscles biopsies. Comparison with controls showed 55 and 46 lncRNAs were differentially expressed in IBM and Jo-1 myositis, respectively. Of these, 16 lncRNAs were differentially expressed in both IBM and Jo-1 myositis and included upregulated H19, lncMyoD and MALAT1. Given that these are known to regulate muscle proliferation and differentiation, we speculate that changes in lncRNAs might contribute to the phenotypic changes in Jo-1 and IBM myositis

New insights into the genetic etiology of Alzheimer's disease and related dementias

Characterization of the genetic landscape of Alzheimer's disease (AD) and related dementias (ADD) provides a unique opportunity for a better understanding of the associated pathophysiological processes. We performed a two-stage genome-wide association study totaling 111,326 clinically diagnosed/'proxy' AD cases and 677,663 controls. We found 75 risk loci, of which 42 were new at the time of analysis. Pathway enrichment analyses confirmed the involvement of amyloid/tau pathways and highlighted microglia implication. Gene prioritization in the new loci identified 31 genes that were suggestive of new genetically associated processes, including the tumor necrosis factor alpha pathway through the linear ubiquitin chain assembly complex. We also built a new genetic risk score associated with the risk of future AD/dementia or progression from mild cognitive impairment to AD/dementia. The improvement in prediction led to a 1.6- to 1.9-fold increase in AD risk from the lowest to the highest decile, in addition to effects of age and the APOE ε4 allele