Weber B, Kimhi S, Howared G, Eden A, Lyko F.. Demethylation of a LINE-1 antisense promoter in the cMet locus impairs Met signaling through induction of illegitimate transcription. Oncogene 29: 5775-5784

Division of Epigenetics, DKFZ-ZMBH Alliance, German Cancer Research Center, Heidelberg, Germany.
Oncogene (Impact Factor: 8.46). 10/2010; 29(43):5775-84. DOI: 10.1038/onc.2010.227
Source: PubMed


The cytosine analogues 5-azacytidine and 5-aza-2'-deoxycytidine are currently the most advanced drugs for epigenetic cancer therapy. Both drugs function as DNA methyltransferase (DNMT) inhibitors and lead to the reactivation of epigenetically silenced tumour suppressor genes. However, not much is known about their target sequence specificity and their possible side effects on normally methylated sequences such as long interspersed nuclear element (LINE)-1 retroelements. It has been shown that demethylation and activation of the LINE-1 antisense promoter can drive the transcription of neighbouring sequences. In this study, we show that demethylation of the colon carcinoma cell line HCT116, either by treatment with DNMT inhibitors or by genetic disruption of the major DNMTs, induces the expression of an illegitimate fusion transcript between an intronic LINE-1 element and the proto-oncogene cMet (L1-cMet). Similar findings were also obtained with myeloid leukaemia cells, an established cellular model for the approved indication of azacytidine and decitabine. Interestingly, upregulation of L1-cMet transcription resulted in reduced cMet expression, which in turn led to decreased cMet receptor signalling. Our results thus provide an important paradigm for demethylation-dependent modulation of gene expression, even if the promoter of the corresponding gene is unmethylated.

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Available from: Amir Eden, Oct 03, 2015
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    • "Previous studies have also suggested the possibility that the effect of the L1 insertion may be derived through the action of the ASP promoter (Speek 2001; Nigumann et al. 2002; Weber et al. 2010). Interestingly, it has been shown that an L1 fusion gene transcribed from the ASP may result in a decrease in the nearby gene expression levels (Weber et al. 2010). The L1 ASP is located 0.6 Mb upstream of PRLR coding region and is in the same orientation, which enables the effect of the ASP on PRLR expression. "
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    ABSTRACT: The immotile short tail sperm (ISTS) defect was recognized in the Finnish Yorkshire population at the end of the 1990s when several affected boars were identified. The causal mutation for this defect is a recent L1 insertion within the SPEF2 gene. In 2001, the insertion frequency was already 0.23. Even though all homozygous boars are eliminated from the population due to infertility, the amount of affected boars increased rapidly until marker-assisted selection against the defect was established. Previously we identified an association between the L1 insertion and litter size in the first parity. In this study, we analyzed the expression of the genomic region adjacent to the L1 insertion on porcine chromosome 16. Based on the RNA-seq data analysis, prolactin receptor (PRLR) was identified as down-regulated in the oviduct of ISTS homozygous sows. Quantitative PCR (qPCR) analysis confirmed the significant down-regulation of PRLR in the ovary, oviduct, and uterus of ISTS homozygous and carrier sows compared with controls. In addition, three unannotated loci between PRLR and SPEF2 showed some transcription activity in the analyzed samples. We further investigated the possible mechanisms of the L1 influence on the decrease in the identified gene expression. The methylation pattern of the PRLR gene region appeared unaffected. However, reads mapping to the L1 sequence indicated an increase in L1 antisense promoter expression in the ISTS homozygous animals. The current data suggest that the presence of the L1 affects by some mechanism the expression patterns upstream of the insertion site.
    Animal Genetics 04/2014; 45(4). DOI:10.1111/age.12153 · 2.21 Impact Factor
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    • "Mouse endogenous retroviruses have been shown to disrupt overlapping gene expression (28,29). Human L1s may affect expression of overlapping genes, including the Met proto-oncogene (30) and others (31). "
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    ABSTRACT: Between 6 and 30% of human and mouse transcripts are initiated from transposable elements. However, the promoters driving such transcriptional activity are mostly unknown. We experimentally characterized an antisense (AS) promoter in mouse L1 retrotransposons for the first time, oriented antiparallel to the coding strand of L1 open reading frame-1. We found that AS transcription is mediated by RNA polymerase II. Rapid amplification of cDNA ends cloning mapped transcription start sites adjacent to the AS promoter. We identified >100 novel fusion transcripts, of which many were conserved across divergent mouse lineages, suggesting conservation of potential functions. To evaluate whether AS L1 transcription could regulate L1 retrotransposition, we replaced portions of native open reading frame-1 in donor elements by synonymously recoded sequences. The resulting L1 elements lacked AS promoter activity and retrotransposed more frequently than endogenous L1s. Overexpression of AS L1 transcripts also reduced L1 retrotransposition. This suppression of retrotransposition was largely independent of Dicer. Our experiments shed new light on how AS fusion transcripts are initiated from endogenous L1 elements across the mouse genome. Such AS transcription can contribute substantially both to natural transcriptional variation and to endogenous regulation of L1 retrotransposition.
    Nucleic Acids Research 02/2014; 42(7). DOI:10.1093/nar/gku091 · 9.11 Impact Factor
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    • "Recent evidence suggests the existence of a causal link between aberrant activation of individual L1-ASP promoters and cancer development and progression. For example, the hypomethylated L1-ASP of an intronic LINE-1 has been shown to act as an alternative promoter driving expression of a truncated isoform of the oncogene cMET (28–30). Our group has also shown that hypomethylation-induced activation of L1-ASP promoters can drive transcription of cancer-specific LCTs transcribed in the same (sense) or opposite (antisense) orientation with respect to neighbouring genes (22). "
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    ABSTRACT: LINE-1 retrotransposons are abundant repetitive elements of viral origin, which in normal cells are kept quiescent through epigenetic mechanisms. Activation of LINE-1 occurs frequently in cancer and can enable LINE-1 mobilization but also has retrotransposition-independent consequences. We previously reported that in cancer, aberrantly active LINE-1 promoters can drive transcription of flanking unique sequences giving rise to LINE-1 chimeric transcripts (LCTs). Here, we show that one such LCT, LCT13, is a large transcript (>300 kb) running antisense to the metastasis-suppressor gene TFPI-2. We have modelled antisense RNA expression at TFPI-2 in transgenic mouse embryonic stem (ES) cells and demonstrate that antisense RNA induces silencing and deposition of repressive histone modifications implying a causal link. Consistent with this, LCT13 expression in breast and colon cancer cell lines is associated with silencing and repressive chromatin at TFPI-2. Furthermore, we detected LCT13 transcripts in 56% of colorectal tumours exhibiting reduced TFPI-2 expression. Our findings implicate activation of LINE-1 elements in subsequent epigenetic remodelling of surrounding genes, thus hinting a novel retrotransposition-independent role for LINE-1 elements in malignancy.
    Nucleic Acids Research 05/2013; 41(14). DOI:10.1093/nar/gkt438 · 9.11 Impact Factor
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