Abstract
Glioblastomas show heterogeneous histological features, and tumor cells show distinct phenotypic states that confer different functional attributes and an aggressive character. However, the molecular mechanisms underlying the heterogeneity in this disease are poorly understood. Glioma stem-like cells (GSCs) are considered able to aberrantly differentiate into diverse cell types and may contribute to the establishment of tumor heterogeneity. Using a GSC model, we investigated differentially expressed microRNAs (miRNAs) and associated epigenetic mechanisms that regulate the differentiation of GSCs. miRNA profiling using microarray technology showed that 13 and 34 miRNAs were commonly up-regulated and down-regulated in two independent GSC lines during differentiation, respectively. Among this set of miRNAs, quantitative PCR analysis showed that miRNA-1275 (miR-1275) was consistently down-regulated during GSC differentiation, along with the up-regulation of its target, CLDN11, an important protein during oligodendroglial lineage differentiation. Inhibition of miR-1275 with a specific antisense oligonucleotide (anti-miR-1275) in GSCs increased the expression of CLDN11, together with significant growth suppression. Epigenetic analysis revealed that gain of histone H3 lysine 27 trimethylation (H3K27me3) in the primary microRNA-1275 promoter was closely associated with miR-1275 expression. Treatment with 3-deazaneplanocin A, an inhibitor of H3K27 methyltransferase, attenuated CLDN11 induction by serum stimulation in parallel with sustained miR-1275 expression. Our results have illuminated the epigenetic regulatory pathways of miR-1275 that are closely associated with oligodendroglial differentiation, which may contribute to the tissue heterogeneity seen in the formation of glioblastomas. Given that inhibition of miR-1275 induces expression of oligodendroglial lineage proteins and suppresses tumor cell proliferation, this may be a potential therapeutic target for glioblastomas.
Highlights
Molecular mechanisms underlying heterogeneity of glioblastoma are poorly understood
Using a Glioma stem-like cells (GSCs) model, we investigated differentially expressed microRNAs and associated epigenetic mechanisms that regulate the differentiation of GSCs. miRNA profiling using microarray technology showed that 13 and 34 miRNAs were commonly up-regulated and down-regulated in two independent GSC lines during differentiation, respectively
To clarify whether these CLDN11-positive cells correspond to the oligodendroglial lineage, which sometimes present in a small fraction of glioblastomas [39, 40], we investigated the expression pattern of CLDN11 in comparison with major neural cell lineage markers, such as glial fibrillary acidic protein (GFAP), microtubule-associated protein 2 (MAP2, a neuron lineage-specific marker), and oligodendrocyte lineage transcription factor 2 (OLIG2, a oligodendrocyte lineage-specific marker)
Summary
Molecular mechanisms underlying heterogeneity of glioblastoma are poorly understood. Results: Newly identified microRNA-1275, which is controlled by a polycomb-mediated silencing mechanism, regulates expression of oligodendroglial lineage protein, Claudin, in glioma stem-like cells. Using a GSC model, we investigated differentially expressed microRNAs (miRNAs) and associated epigenetic mechanisms that regulate the differentiation of GSCs. miRNA profiling using microarray technology showed that 13 and 34 miRNAs were commonly up-regulated and down-regulated in two independent GSC lines during differentiation, respectively Among this set of miRNAs, quantitative PCR analysis showed that miRNA1275 (miR-1275) was consistently down-regulated during GSC differentiation, along with the up-regulation of its target, CLDN11, an important protein during oligodendroglial lineage differentiation. We investigated the molecular effects of the newly identified miR-1275 in GSC differentiation, whose expression is regulated via a PRC2-H3K27me3-dependent epigenetic mechanism in response to environmental cues, and assessed the relationship between stem-like cell differentiation and tissue heterogeneity, especially the oligodendroglial component in glioblastomas. Our results suggest that this developmental microRNA is regulated via an epigenetic pathway that contributes to the phenotypic diversity of glioblastoma tissues, which may in turn provide a better understanding of the heterogeneity of glioblastoma in the context of human neurodevelopment
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