Decoupling of DNA methylation and activity of intergenic LINE-1 promoters in colorectal cancer

Natasha Vafadar-Isfahani,Christina Parr,Lara E Mcmillan,Juliane Sanner,Zhao Yeo,Stephen Saddington,Oliver Peacock,Hazel A Cruickshanks,Richard R Meehan,Jonathan N Lund,Cristina Tufarelli

doi:10.1080/15592294.2017.1300729

Natasha Vafadar-Isfahani, Christina Parr + Show 9 more

Open Access

https://doi.org/10.1080/15592294.2017.1300729

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Abstract

ABSTRACTHypomethylation of LINE-1 repeats in cancer has been proposed as the main mechanism behind their activation; this assumption, however, was based on findings from early studies that were biased toward young and transpositionally active elements. Here, we investigate the relationship between methylation of 2 intergenic, transpositionally inactive LINE-1 elements and expression of the LINE-1 chimeric transcript (LCT) 13 and LCT14 driven by their antisense promoters (L1-ASP). Our data from DNA modification, expression, and 5′RACE analyses suggest that colorectal cancer methylation in the regions analyzed is not always associated with LCT repression. Consistent with this, in HCT116 colorectal cancer cells lacking DNA methyltransferases DNMT1 or DNMT3B, LCT13 expression decreases, while cells lacking both DNMTs or treated with the DNMT inhibitor 5-azacytidine (5-aza) show no change in LCT13 expression. Interestingly, levels of the H4K20me3 histone modification are inversely associated with LCT13 and LCT14 expression. Moreover, at these LINE-1s, H4K20me3 levels rather than DNA methylation seem to be good predictor of their sensitivity to 5-aza treatment. Therefore, by studying individual LINE-1 promoters we have shown that in some cases these promoters can be active without losing methylation; in addition, we provide evidence that other factors (e.g., H4K20me3 levels) play prominent roles in their regulation.

Highlights

Long interspersed element 1 (LINE-1) regions are repetitive DNA sequences that comprise about 17% of the human genome, equivalent to approximately eight times the protein coding portion of the genome [1]
We have investigated at a locusspecific level if loss of DNA methylation within the L1-5’UTR of the LINE-1s driving LCT13 and LCT14 is always associated with their expression, and we have compared the presence of histone marks associated with active and inactive chromatin states to transcriptional activity and DNA methylation levels at these sites
Three microsatellite instable (MSI: HCA-7, HCT116, RKO) and three microsatellite stable (MSS: CaCo-2; SW480 and SW620) colorectal cancer (CRC) cell lines were tested as higher levels of methylation at LINE-1s have been reported in MSI relatively to MSS lines [36]

Summary

Introduction

Long interspersed element 1 (LINE-1) regions are repetitive DNA sequences that comprise about 17% of the human genome, equivalent to approximately eight times the protein coding portion of the genome [1]. Of the about 516,000 LINE-1s present in the human genome, just around 7000 have retained potentially intact promoters; of these, nearly 5000 are full-length but only up to 20 so called ‘hot-L1s’ have been shown to be able to retrotranspose [3,4,5]. It is becoming more widely accepted that LINE-1s can contribute to tumor in ways that are not related to their mobilization [6,7,8]. Transcription initiation from L1-ASP in cancer has been found between position 160 and 200 of the L1-5’UTR, in particular for intergenic, retrotransposition deficient LINE-1s [10,15]

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