Over-expressed lncRNA HOTAIRM1 promotes tumor growth and invasion through up-regulating HOXA1 and sequestering G9a/EZH2/Dnmts away from the HOXA1 gene in glioblastoma multiforme

Abstract

Background

Glioblastoma multiforme (GBM) is the common primary brain tumor classified the most malignant glioma. Long non-coding RNAs (LncRNAs) are important epigenetic regulators with critical roles in cancer initiation and progression. LncRNA HOTAIRM1 transcribes from the antisense strand of HOXA gene cluster which locus in chromosome 7p15.2. Recent studies have shown that HOTAIRM1 is involved in acute myeloid leukemia and colorectal cancer. Here we sought to investigate the role of HOTAIRM1 in GBM and explore its mechanisms of action.

Methods

The expressions of HOTAIRM1 and HOXA1 in GBM tissues and cells were determined by qRT-PCR, and the association between HOTAIRM1, HOXA1 transcription and tumor grade were analyzed. The biological function of HOTAIRM1 in GBM was evaluated both in vitro and in vivo. Chromatin immunoprecipitation (ChIP) assay and quantitative Sequenom MassARRAY methylation analysis were performed to explore whether HOTAIRM1 could regulate histone and DNA modification status of the HOXA1 gene transcription start sites (TSS) and activate its transcription. ChIP and RNA-ChIP were further performed to determine the molecular mechanism of HOTAIRM1 in epigenetic regulation of the HOXA1 gene.

Results

HOTAIRM1 was abnormally up-regulated in GBM tissues and cells, and this up-regulation was correlated with grade malignancy in glioma patients. HOTAIRM1 silencing caused tumor suppressive effects via inhibiting cell proliferation, migration and invasion, and inducing cell apoptosis. In vivo experiments showed knockdown of HOTAIRM1 lessened the tumor growth. Additionally, HOTAIRM1 action as regulating the expression of the HOXA1 gene. HOXA1, as an oncogene, it’s expression levels were markedly elevated in GBM tissues and cell lines. Mechanistically, HOTAIRM1 mediated demethylation of histone H3K9 and H3K27 and reduced DNA methylation levels by sequester epigenetic modifiers G9a and EZH2, which are H3K9me² and H3K27me³ specific histone methyltransferases, and DNA methyltransferases (DnmTs) away from the TSS of HOXA1 gene.

Conclusions

We investigated the potential role of HOTAIRM1 to promote GBM cell proliferation, migration, invasion and inhibit cell apoptosis by epigenetic regulation of HOXA1 gene that can be targeted simultaneously to effectively treat GBM, thus putting forward a promising strategy for GBM treatment. Meanwhile, this finding provides an example of transcriptional control over the chromatin state of gene and may help explain the role of lncRNAs within the HOXA gene cluster.