Attenuated adenosine-to-inosine editing of microRNA-376a* promotes invasiveness of glioblastoma cells
Yukti Choudhury, … , Beng-Ti Ang, Shu Wang
Yukti Choudhury, … , Beng-Ti Ang, Shu Wang
Published October 24, 2012
Citation Information: J Clin Invest. 2012;122(11):4059-4076. https://doi.org/10.1172/JCI62925.
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Research Article

Attenuated adenosine-to-inosine editing of microRNA-376a* promotes invasiveness of glioblastoma cells

Abstract

In the human brain, microRNAs (miRNAs) from the microRNA-376 (miR-376) cluster undergo programmed “seed” sequence modifications by adenosine-to-inosine (A-to-I) editing. Emerging evidence suggests a link between impaired A-to-I editing and cancer, particularly in high-grade gliomas. We hypothesized that disruption of A-to-I editing alters expression of genes regulating glioma tumor phenotypes. By sequencing the miR-376 cluster, we show that the overall miRNA editing frequencies were reduced in human gliomas. Specifically in high-grade gliomas, miR-376a* accumulated entirely in an unedited form. Clinically, a significant correlation was found between accumulation of unedited miR-376a* and the extent of invasive tumor spread as measured by magnetic resonance imaging of patient brains. Using both in vitro and orthotopic xenograft mouse models, we demonstrated that the unedited miR-376a* promoted glioma cell migration and invasion, while the edited miR-376a* suppressed these features. The effects of the unedited miR-376a* were mediated by its sequence-dependent ability to target RAP2A and concomitant inability to target AMFR. Thus, the tumor-dependent introduction of a single base difference in the miR-376a* sequence dramatically alters the selection of its target genes and redirects its function from inhibiting to promoting glioma cell invasion. These findings uncover a new mechanism of miRNA deregulation and identify unedited miR-376a* as a potential therapeutic target in glioblastoma cells.

Authors

Yukti Choudhury, Felix Chang Tay, Dang Hoang Lam, Edwin Sandanaraj, Carol Tang, Beng-Ti Ang, Shu Wang

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Figure 1

A-to-I editing of the miR-376 cluster is reduced in gliomas.

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A-to-I editing of the miR-376 cluster is reduced in gliomas. (A) Primary...
(A) Primary miRNAs encoded by the human miR-376 cluster. Mature miRNAs processed from the pri-miRNAs are indicated in green. Positions of editing sites are indicated by red arrows. (B) Chromatograms from direct sequencing of RT-PCR products of pri-miR-376a1 from normal human brain (NB), GBM (GBM 1), and AA (AA 20). The genomically encoded sequence is shown. Underlined regions correspond to two mature miRNAs, miR-376a* and miR-376a, processed from pri-miR-376a1. Editing sites are indicated by red arrows. (C) Editing frequency at the +9 site of pri-miR-376a1 (miR-376a*) in samples of NB (n = 5), GBM (n = 12), AA (n = 4), and AOAs and AOGs (n = 4). Means are designated by horizontal lines. (D) Editing frequency of mature miRNAs from the miR-376 cluster in glioma cell lines. cDNA clones (n = 50) corresponding to mature miRNA were sequenced, and frequency of editing was calculated as the percentage of clones with A-to-G change at the editing site. (E) Direct sequencing of RT-PCR products of pri-miR-376a1 from U87 cells co-transfected with a plasmid expressing ADARB1 and the human miR-376 cluster. Red arrows indicate editing sites. Site corresponding to miR-376a* is boxed. A plasmid expressing EGFP was used as a control. (F) Relative expression of ADARB1 mRNA, by qRT-PCR in NB and tumor samples. Data represent mean ± SD (n = 3).