The Intronic Long Noncoding RNA ANRASSF1 Recruits PRC2 to the RASSF1A Promoter, Reducing the Expression of RASSF1A and Increasing Cell Proliferation

Abstract

The down-regulation of the tumor-suppressor gene RASSF1A has been shown to increase cell proliferation in several tumors. RASSF1A expression is regulated through epigenetic events involving the polycomb repressive complex 2 (PRC2); however, the molecular mechanisms modulating the recruitment of this epigenetic modifier to the RASSF1 locus remain largely unknown. Here, we identify and characterize ANRASSF1, an endogenous unspliced long noncoding RNA (lncRNA) that is transcribed from the opposite strand on the RASSF1 gene locus in several cell lines and tissues and binds PRC2. ANRASSF1 is transcribed through RNA polymerase II and is 5′-capped and polyadenylated; it exhibits nuclear localization and has a shorter half-life compared with other lncRNAs that bind PRC2. ANRASSF1 endogenous expression is higher in breast and prostate tumor cell lines compared with non-tumor, and an opposite pattern is observed for RASSF1A. ANRASSF1 ectopic overexpression reduces RASSF1A abundance and increases the proliferation of HeLa cells, whereas ANRASSF1 silencing causes the opposite effects. These changes in ANRASSF1 levels do not affect the RASSF1C isoform abundance. ANRASSF1 overexpression causes a marked increase in both PRC2 occupancy and histone H3K27me3 repressive marks, specifically at the RASSF1A promoter region. No effect of ANRASSF1 overexpression was detected on PRC2 occupancy and histone H3K27me3 at the promoter regions of RASSF1C and the four other neighboring genes, including two well-characterized tumor suppressor genes. Additionally, we demonstrated that ANRASSF1 forms an RNA/DNA hybrid and recruits PRC2 to the RASSF1A promoter. Together, these results demonstrate a novel mechanism of epigenetic repression of the RASSF1A tumor suppressor gene involving antisense unspliced lncRNA, in which ANRASSF1 selectively represses the expression of the RASSF1 isoform overlapping the antisense transcript in a location-specific manner. In a broader perspective, our findings suggest that other non-characterized unspliced intronic lncRNAs transcribed in the human genome might contribute to a location-specific epigenetic modulation of genes.

Long Noncoding RNA ANRIL Promotes Non–Small Cell Lung Cancer Cell Proliferation and Inhibits Apoptosis by Silencing KLF2 and P21 Expression

“…These findings suggest that ANRIL plays a direct role in the modulation of cell proliferation and NSCLC progression, and could be a useful novel prognostic or progression marker for NSCLC. As more and more lncRNAs are studied, many have been shown to function by binding with PRC2 and silencing downstream target genes that involved in multiple cancers, including NSCLC (26,27). ANRIL has been reported to involve in cancer cells proliferation by silencing p15 INK4 expression.…”

RASSF tumor suppressor gene family: Biological functions and regulation

“…Another player involved in epigenetic regulation of RASSF1A was reported recently [154]. Beckedorff et al showed that nonspliced long non-coding RNA transcribed from the antisense strand of RASSF1A forms an RNA/DNA hybrid at the RASSF1A transcription site and recruits polycomb repressor complex 2 (PRC2) to the RASS-F1A promoter.…”

Long Noncoding RNA PVT1 Promotes Non–Small Cell Lung Cancer Cell Proliferation through Epigenetically Regulating LATS2 Expression

“…For example, RASSF1 antisense 1 (ANRASSF1) recruits PRC2 to the RASSF1A promoter, resulting in silenced expression of RASSF1A and increased cell proliferation. Ubiquitin-conjugating enzyme E2K (UBE2K) acts as a negative prognostic factor for lymph node metastasis and survival via associating with PRC2 in bladder cancer (15,16).…”