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MicroRNA-96 directly inhibits ?-globin expression in human erythropoiesis.

ABSTRACT: Fetal hemoglobin, HbF (?(2)?(2)), is the main hemoglobin synthesized up to birth, but it subsequently declines and adult hemoglobin, HbA (?(2)?(2)), becomes predominant. Several studies have indicated that expression of the HbF subunit ?-globin might be regulated post-transcriptionally. This could be confered by ?22-nucleotide long microRNAs that associate with argonaute proteins to specifically target ?-globin mRNAs and inhibit protein expression. Indeed, applying immunopurifications, we found that ?-globin mRNA was associated with argonaute 2 isolated from reticulocytes that contain low levels of HbF (<1%), whereas association was significantly lower in reticulocytes with high levels of HbF (90%). Comparing microRNA expression in reticulocytes from cord blood and adult blood, we identified several miRNAs that were preferentially expressed in adults, among them miRNA-96. The overexpression of microRNA-96 in human ex vivo erythropoiesis decreased ?-globin expression by 50%, whereas the knock-down of endogenous microRNA-96 increased ?-globin expression by 20%. Moreover, luciferase reporter assays showed that microRNA-96 negatively regulates expression of ?-globin in HEK293 cells, which depends on a seedless but highly complementary target site located within the coding sequence of ?-globin. Based on these results we conclude that microRNA-96 directly suppresses ?-globin expression and thus contributes to HbF regulation.

SUBMITTER: Azzouzi I

PROVIDER: S-EPMC3145767 | biostudies-literature | 2011

REPOSITORIES: biostudies-literature

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MicroRNA-96 directly inhibits γ-globin expression in human erythropoiesis.

Azzouzi Imane I Moest Hansjoerg H Winkler Jeannine J Fauchère Jean-Claude JC Gerber André P AP Wollscheid Bernd B Stoffel Markus M Schmugge Markus M Speer Oliver O

PloS one 20110728 7

Fetal hemoglobin, HbF (α(2)γ(2)), is the main hemoglobin synthesized up to birth, but it subsequently declines and adult hemoglobin, HbA (α(2)β(2)), becomes predominant. Several studies have indicated that expression of the HbF subunit γ-globin might be regulated post-transcriptionally. This could be confered by ∼22-nucleotide long microRNAs that associate with argonaute proteins to specifically target γ-globin mRNAs and inhibit protein expression. Indeed, applying immunopurifications, we found ...[more]

PMID: 21829531

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Project description:Glioma is one of the most common, rapidly progressive and fatal brain tumors, and accumulating evidence shows that microRNAs (miRNAs) play important roles in the development of cancers, including glioma. Therapeutic applications of miRNAs in Ras-driven glioma have been proposed; however, their specific functions and mechanisms are poorly understood. Here, we report that miR-1301-3p directly targets the neuroblastoma Ras viral oncogene homolog (N-Ras) and functions as a tumor-suppressor in glioma. Quantitative reverse transcription-PCR was applied to detect the expression of miR-1301-3p in glioma specimens. The direct target genes of miR-1301-3p were predicted by bioinformatic analysis and further verified by immunoblotting and luciferase assays. The effects of miR-1301-3p on the proliferation and cell cycle of glioma cells were analyzed by cell-counting kit 8, colony formation, 5-ethynyl-2-deoxyuridine (EDU) and flow cytometry assays. A xenograft model was used to study the effect of miR-1301-3p on tumor growth and angiogenesis. The expression levels of miR-1301-3p in glioma specimens were significantly downregulated. N-Ras was confirmed as a direct target of miR-1301-3p. MiR-1301-3p inhibited glioma cell growth and blocked the cell cycle to G1 by negatively regulating N-Ras and its downstream signaling pathway, MEK-ERK1/2. Furthermore, the inhibitory effects of miR-1301-3p could be rescued by the overexpression of N-Ras. The protein levels of N-Ras were up-regulated in clinical glioma specimens and were negatively correlated with miR-1301-3p expression levels (r=-056, P=0.0002). In vivo studies revealed that increased levels of miR-1301-3p delayed the growth of intracranial tumors, which was accompanied by decreased Ki67 and CD31 expression. Taken together, our results demonstrate that miR-1301-3p plays a significant role in inactivating the Ras signaling pathway through the inhibition of N-Ras, which may provide a novel therapeutic strategy for treatment of glioma and other Ras-driven cancers.

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MicroRNA-96 directly inhibits ?-globin expression in human erythropoiesis.

Publications

MicroRNA-96 directly inhibits γ-globin expression in human erythropoiesis.

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