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Chromosome Interactions at Promoters Reveal Non-Coding Colorectal Cancer Mutations

NEW YORK (GenomeWeb) – A team led by researchers at the Institute of Cancer Research has used chromosome conformation capture (Hi-C) to help pinpoint recurrent, non-coding mutations related to cancer development in the colorectal (CRC) cancer genome.

"This study reveals new insights into the complex genetic alterations driving tumor development, providing a paradigm for employing chromosome conformation capture to decipher non-coding CREs relevant to cancer biology," corresponding author Richard Houlston, a genetics and epidemiology researcher at ICR, and his colleagues wrote today in Nature Genetics.

The researchers relied on high-throughput capture Hi-C (CHi-C) to assess 19,023 promoter fragment sites in the microsatellite stable HT29 CRC cell line and the LoVo cell line, representing microsatellite instable CRC. They searched for informative cis-regulatory element (CREs) interactions in microsatellite-stable and -instable CRC cell lines. They reasoned that "CREs that modulate gene expression represent a highly enriched subset of the non-coding genome in which to search for driver mutations."

Along with corresponding whole-genome sequencing, RNA sequence-based gene expression profiles, and copy number data generated for the Cancer Genome Atlas project, the CHi-C profiles led them to a recurrent regulatory mutation in the ETV1 promoter in microsatellite stable tumors. In the process, they tracked down almost 118,800 contacts involving promoter fragments from the HT29 line and nearly 96,500 promoter fragment contacts from the LoVo line.

The team's subsequent analysis suggested that the ETV1 promoter change can influence the expression of the proposed oncogene. And the analysis uncovered other regulatory interactions at mutation-prone sites as well, including copy number variants that affect regulatory regions of the RASL11A gene.

By combining the CHi-C profiles with genomic information for colon and rectal adenocarcinoma tumors profiled for TCGA — including whole genome sequence data for dozens of colon and rectal cancers and hundreds more CRCs assessed by Affymetrix SNP arrays — the researchers went on to determine whether CHi-C interactions fell near active or inactive genes, explore the recurrent single nucleotide or copy number changes falling at these sites, search for signs of positive selection, and retrace the transcriptional consequences of such non-coding alterations.

When the researchers used CRISPR-Cas9 gene editing to follow up on the RASL11A promoter site in cancer cell lines, for example, they found that deletions in the promoter could dial down RASL11A expression, while amplifications affecting the CRE were linked to enhanced RASL11A expression.

"[O]ur work supports the existence of non-coding drivers for CRC, and more broadly provides a paradigm for using chromosome conformation capture to decode disease-specific regulatory elements," the authors concluded. "Such discoveries facilitate the identification of novel therapeutic and chemoprevention agents, and classification of patients into molecular subgroups to personalize therapy."