RESUMO
BACKGROUND: ADAR enzymes convert adenosines to inosines within double-stranded RNAs, including microRNA (miRNA) precursors, with important consequences on miRNA retargeting and expression. ADAR2 activity is impaired in glioblastoma and its rescue has anti-tumoral effects. However, how ADAR2 activity may impact the miRNome and the progression of glioblastoma is not known. RESULTS: By integrating deep-sequencing and array approaches with bioinformatics analyses and molecular studies, we show that ADAR2 is essential to edit a small number of mature miRNAs and to significantly modulate the expression of about 90 miRNAs in glioblastoma cells. Specifically, the rescue of ADAR2 activity in cancer cells recovers the edited miRNA population lost in glioblastoma cell lines and tissues, and rebalances expression of onco-miRNAs and tumor suppressor miRNAs to the levels observed in normal human brain. We report that the major effect of ADAR2 is to reduce the expression of a large number of miRNAs, most of which act as onco-miRNAs. ADAR2 can edit miR-222/221 and miR-21 precursors and decrease the expression of the corresponding mature onco-miRNAs in vivo and in vitro, with important effects on cell proliferation and migration. CONCLUSIONS: Our findings disclose an additional layer of complexity in miRNome regulation and provide information to better understand the impact of ADAR2 editing enzyme in glioblastoma. We propose that ADAR2 is a key factor for maintaining edited-miRNA population and balancing the expression of several essential miRNAs involved in cancer.
Assuntos
Adenosina Desaminase/metabolismo , Neoplasias Encefálicas/genética , Regulação Neoplásica da Expressão Gênica , Glioblastoma/enzimologia , Glioblastoma/genética , MicroRNAs/genética , Edição de RNA/genética , Proteínas de Ligação a RNA/metabolismo , Adolescente , Animais , Encéfalo/enzimologia , Neoplasias Encefálicas/enzimologia , Neoplasias Encefálicas/patologia , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Regulação para Baixo/genética , Perfilação da Expressão Gênica , Inativação Gênica , Glioblastoma/patologia , Células HEK293 , Humanos , Camundongos , MicroRNAs/metabolismo , Modelos BiológicosRESUMO
MicroRNAs (miRNAs) represent a class of small non-coding RNAs that control gene expression by targeting mRNAs and triggering either repression of translation or RNA degradation. They have been shown to be involved in a variety of biological processes such as development, differentiation and cell cycle control, but little is known about their involvement in the response to irradiation. We showed here that in human umbilical vein endothelial cells (HUVEC) some miRNAs previously shown to have a crucial role in vascular biology are transiently modulated in response to a clinically relevant dose of ionizing radiation. In particular we identified an early transcriptional induction of several members of the microRNA cluster 17-92 and other microRNAs already known to be related to angiogenesis. At the same time we observed a peculiar behavior of the miR-221/222 cluster, suggesting an important role of these microRNAs in HUVEC homeostasis. We observed an increased efficiency in the formation of capillary-like structures in irradiated HUVEC. These results could lead to a new interpretation of the effect of ionizing radiation on endothelial cells and on the response of tumor endothelial bed cells to radiotherapy.
Assuntos
Células Endoteliais/metabolismo , Células Endoteliais/efeitos da radiação , Regulação da Expressão Gênica/efeitos da radiação , MicroRNAs/genética , Neovascularização Fisiológica/genética , Neovascularização Fisiológica/efeitos da radiação , Cordão Umbilical/citologia , Regiões 3' não Traduzidas/genética , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/genética , Sítios de Ligação , Capilares/citologia , Núcleo Celular/metabolismo , Núcleo Celular/efeitos da radiação , Relação Dose-Resposta à Radiação , Regulação da Expressão Gênica/genética , Células HeLa , Humanos , Transferência Linear de Energia , Estresse Oxidativo/genética , Estresse Oxidativo/efeitos da radiação , Proteínas Proto-Oncogênicas c-myc/metabolismo , Raios XRESUMO
OBJECTIVE: Smad-interacting protein-1 (Sip1/ZEB2) is a transcriptional repressor of the telomerase reverse transcriptase catalytic subunit (Tert) and has recently been identified as a key regulator of embryonic cell fate with a phenotypic effect similar, in our opinion, to that reported for nitric oxide (NO). Remarkably, SIP1/ZEB2 is a known target of the microRNA 200 (miR-200) family. In this light, we postulated that Sip1/ZEB2 and the miR-200 family could play a role during the NO-dependent differentiation of mES. METHODS AND RESULTS: The results of the present study show that Sip1/ZEB2 expression is downregulated during the NO-dependent expression of mesendoderm and early cardiovascular precursor markers, including Flk1 and CXCR4 in mES. Coincidently, members of the miR-200 family, namely miR-429, -200a, -200b, and -200c, were transcriptionally induced in parallel to mouse Tert. This regulation occurred at the level of chromatin. Remarkably, miR-429/miR-200a overexpression or Sip1/ZEB2 knockdown by short hairpin RNA interference elicited a gene expression pattern similar to that of NO regardless of the presence of leukemia inhibitory factor. CONCLUSIONS: These results are the first demonstrating that the miR-200 family and Sip1/ZEB2 transcription factor are regulated by NO, indicating an unprecedented molecular circuitry important for telomerase regulation and early differentiation of mES.
Assuntos
Diferenciação Celular , Células-Tronco Embrionárias/metabolismo , Proteínas de Homeodomínio/metabolismo , MicroRNAs/metabolismo , Óxido Nítrico/metabolismo , Proteínas Repressoras/metabolismo , Transdução de Sinais , Animais , Diferenciação Celular/efeitos dos fármacos , Células Cultivadas , Montagem e Desmontagem da Cromatina , Células-Tronco Embrionárias/efeitos dos fármacos , Regulação da Expressão Gênica no Desenvolvimento , Proteínas de Homeodomínio/genética , Fator Inibidor de Leucemia/metabolismo , Camundongos , Doadores de Óxido Nítrico/farmacologia , Interferência de RNA , RNA Mensageiro/metabolismo , Proteínas Repressoras/genética , Transdução de Sinais/efeitos dos fármacos , Telomerase/metabolismo , Fatores de Tempo , Transcrição Gênica , Transfecção , Homeobox 2 de Ligação a E-box com Dedos de ZincoRESUMO
The B cell lymphomas associated with Epstein-Barr virus (EBV) are not limited to any specific stage of B cell differentiation but covers widely different B cell phenotypes. In vitro infection of the virus negative tumors with a recombinant EBV strain has provided important insights into virus-tumor interaction. Here, we investigated the interaction between EBV and terminally differentiated tumor derived B cells, namely multiple myeloma (MM). The in vitro EBV infected MM expressed restricted viral latency. Acquisition of the virus was accompanied by a partial reprogramming to a mature B cell phenotype. Thus, the plasma cell markers syndecan-1 (CD138), Blimp1 and MUM1 were downregulated, while expression of HLADR, CIITA and TCL1, which are normally not expressed in plasmacytoid cells, was upregulated. The silenced transcription factor gene encoding Pax5 and its target BLNK were activated. Significantly, the free lambda light chains secreted in the medium were reduced in EBV infected MM clones. Collectively, these results suggest that the restricted EBV latency can cause at least partial phenotypic reversion of terminally differentiated B tumor cells. We suggest that the restricted EBV latent gene expression may not only be the consequence but the cause of the mature B cell phenotype, actively participating in the virus persistence.