Small RNA Sequencing Uncovers New miRNAs and moRNAs Differentially Expressed in Normal and Primary Myelofibrosis CD34+ Cells

Paola Guglielmelli,Alessandro Coppe,Carmela Mannarelli,Claudia Saccoman,Andrea Bisognin,Alessandro M Vannucchi,Stefania Bortoluzzi,Fabio Martelli

doi:10.1371/journal.pone.0140445

Paola Guglielmelli, Alessandro Coppe + Show 6 more

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https://doi.org/10.1371/journal.pone.0140445

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Journal: PloS one	Publication Date: Oct 15, 2015
Citations: 21	License type: CC BY 4.0

Affiliation: University of Florence, University of Padua

Abstract

Myeloproliferative neoplasms (MPN) are chronic myeloid cancers thought to arise at the level of CD34+ hematopoietic stem/progenitor cells. They include essential thrombocythemia (ET), polycythemia vera (PV) and primary myelofibrosis (PMF). All can progress to acute leukemia, but PMF carries the worst prognosis. Increasing evidences indicate that deregulation of microRNAs (miRNAs) might plays an important role in hematologic malignancies, including MPN. To attain deeper knowledge of short RNAs (sRNAs) expression pattern in CD34+ cells and of their possible role in mediating post-transcriptional regulation in PMF, we sequenced with Illumina HiSeq2000 technology CD34+ cells from healthy subjects and PMF patients. We detected the expression of 784 known miRNAs, with a prevalence of miRNA up-regulation in PMF samples, and discovered 34 new miRNAs and 99 new miRNA-offset RNAs (moRNAs), in CD34+ cells. Thirty-seven small RNAs were differentially expressed in PMF patients compared with healthy subjects, according to microRNA sequencing data. Five miRNAs (miR-10b-5p, miR-19b-3p, miR-29a-3p, miR-379-5p, and miR-543) were deregulated also in PMF granulocytes. Moreover, 3’-moR-128-2 resulted consistently downregulated in PMF according to RNA-seq and qRT-PCR data both in CD34+ cells and granulocytes. Target predictions of these validated small RNAs de-regulated in PMF and functional enrichment analyses highlighted many interesting pathways involved in tumor development and progression, such as signaling by FGFR and DAP12 and Oncogene Induced Senescence. As a whole, data obtained in this study deepened the knowledge of miRNAs and moRNAs altered expression in PMF CD34+ cells and allowed to identify and validate a specific small RNA profile that distinguishes PMF granulocytes from those of normal subjects. We thus provided new information regarding the possible role of miRNAs and, specifically, of new moRNAs in this disease.

Highlights

Philadelphia-negative chronic myeloproliferative neoplasms (MPNs) are a heterogeneous group of clonal hematopoietic stem cell (HSC) disorders associated with overproduction of mature myeloid cells[1,2].Myeloproliferative neoplasms (MPN) are a group of chronic myeloid cancers that include essential thrombocythemia (ET), polycythemia vera (PV) and primary myelofibrosis (PMF), associated with an increased risk of thrombosis and/or hemorrhage and a propensity to develop acute myeloid leukemia.Myelofibrosis, the most symptomatic and associated with the worst survival, can arise primarily or as a progression of PV or ET
We considered small RAS oncogene family POLR2H - Polymerase (RNA) sequencing data of 6 CD34+ cells, including 3 samples collected from 3 pools of bone marrow CD34+ cells of healthy subjects (CTR), and 3 samples of circulating CD34+ cells of patients affected by primary myelofibrosis (PMF), two of which were from individual patients while the third constituted a pool obtained by mixing equal amounts of RNA from 4 PMF patients
We reported the results of the first RNA-seq project aiming at studying small RNAs in PMF CD34+ cells compared with control CD34+ cells

Summary

Introduction

Philadelphia-negative chronic myeloproliferative neoplasms (MPNs) are a heterogeneous group of clonal hematopoietic stem cell (HSC) disorders associated with overproduction of mature myeloid cells[1,2].MPNs are a group of chronic myeloid cancers that include essential thrombocythemia (ET), polycythemia vera (PV) and primary myelofibrosis (PMF), associated with an increased risk of thrombosis and/or hemorrhage and a propensity to develop acute myeloid leukemia.Myelofibrosis, the most symptomatic and associated with the worst survival, can arise primarily or as a progression of PV or ET (post-PV/ET MF). Philadelphia-negative chronic myeloproliferative neoplasms (MPNs) are a heterogeneous group of clonal hematopoietic stem cell (HSC) disorders associated with overproduction of mature myeloid cells[1,2]. MPNs are a group of chronic myeloid cancers that include essential thrombocythemia (ET), polycythemia vera (PV) and primary myelofibrosis (PMF), associated with an increased risk of thrombosis and/or hemorrhage and a propensity to develop acute myeloid leukemia. In primary myelofibrosis (PMF) the increased proliferation of megakaryocytes is accompanied by deposition of fibrosis in the bone marrow, abnormal stem cell trafficking, and extramedullary hematopoiesis (myeloid metaplasia).[1,2]. In 2005, a new somatic mutation in the Janus Kinase 2 (JAK2)[3,4,5,6,7] has been discovered, providing the first genetic insight into the MPN pathogenesis. Other mutations have been discovered, including mutation in MPL gene or in epigenetic regulators.[2,8,9,10]

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