No effect of cancer-associated SNP rs6983267 in the 8q24 region on co-expression of MYC and TCF7L2 in normal colon tissue
© Prokunina-Olsson and Hall; licensee BioMed Central Ltd. 2009
Received: 23 July 2009
Accepted: 06 November 2009
Published: 06 November 2009
A single nucleotide polymorphism (SNP) rs6983267, located within the 8q24 region, is strongly associated with risk of colorectal and prostate cancer. It has been suggested that the mechanism of this association is related to differential interaction of TCF7L2 protein (previously known as TCF-4) with alleles of rs6983267, influencing the expression of a well-known oncogene, MYC, located 335 Kb telomeric. Here, we tested the correlation between mRNA expression of MYC and several alternatively spliced forms of TCF7L2 in 117 non-cancer colon samples. We observed a strong correlation (r = 0.60, p < 10-6) between expression of MYC and a unique splicing form of TCF7L2. The level of MYC expression in these samples was associated with expression of some TCF7L2 splicing forms but not with genotypes of rs6983267, or interaction of rs6983267 with TCF7L2 expression. These findings suggest that some splicing forms of TCF7L2 may be functionally important for regulation of MYC expression in colon tissue but this regulation is not directly dependent on rs6983267.
Recent genome-wide association studies (GWAS) have identified a single nucleotide polymorphism (SNP) rs6983267 within the 8q24 region associated with increased susceptibility to colorectal and prostate cancer [1–6]. Follow-up association studies have suggested that the same variant may also increase the risk for cancers of the kidney, thyroid and larynx [7, 8]. The location of rs6983267 in the intergenic region 335 Kb upstream from the MYC gene, a well-known oncogene , generated a hypothesis that this SNP might be involved in a long-distance regulation of MYC expression. Located in a region with significant evolutionary conservation and enhancer potential [10–12], the SNP was predicted to affect a binding site for TCF7L2 [10, 11], a key transcription factor in the WNT pathway. The risk allele G of rs6983267 was found to have a slightly stronger affinity to TCF7L2 in binding assays compared to the non-risk allele T, and stronger regulatory activity in luciferase reporter assays [10, 11]. An analysis of long-range interactions showed that the region containing rs6983267 might be in physical proximity with MYC region . These findings suggested that rs6983267 might be located within an enhancer element that interacts with TCF7L2 and regulates MYC expression [10, 11]. MYC is a target gene of TCF7L2 [13–15] and its expression is regulated through two TCF7L2 binding sites within the MYC promoter . No association has been found between rs6983267 and the mRNA expression of MYC in lymphoblastoid cell lines [11, 16], normal and tumor colon samples [10, 11, 17–20], or with MYC immunostaining in colon tumors .
Previously, we performed a detailed study of TCF7L2 expression in several types of human tissue, including colon where we measured the expression of multiple assays targeting the majority of known splicing forms of TCF7L2 [21, 22]. In the current study we sought to determine, whether the expression of TCF7L2 splicing forms we identified in non-cancer colon samples correlated with MYC expression and whether this expression was dependent on alleles of rs6983267 or interaction of rs6983267 with TCF7L2 expression.
Correlation between expression of MYC and TCF7L2 in colon samples
MYC assay 1
MYC assay 2
2.4 × 10-5
2.1 × 10-4
1.2 × 10-5
2.5 × 10-4
Association of rs6983267 with MYC expression in colon tissue
MYC assay 1, p-value *
MYC assay 2, p-value *
Our results show a strong role of TCF7L2 in regulation of MYC expression in colon, but not through rs6983267. Both TCF7L2 and MYC genes are expressed in colon and are important for maintaining proliferation of intestinal epithelium [28, 29] (Additional file 3). Inactivation of the adenomatous polyposis coli (APC) tumor suppressor gene leads to formation of β-catenin/TCF7L2 complexes, constitutive activation of the WNT pathway and eventually colorectal cancer. Rare point mutations within TCF7L2 are also found in colorectal cancers . The proliferative effect of TCF7L2 is achieved through its transcriptional regulation of several target genes such as MYC and CCND1 (Cyclin D1). Our results suggest that expression of MYC in colon tissue is most likely regulated by a splicing form of TCF7L2 encoding a protein with a potent transactivation CRARF-domain. However, we did not find any evidence for an effect of rs6983267 on TCF7L2 regulation of MYC expression. Of the family of TCF/LEF transcription factors, TCF7L2 has the highest expression in colon, but other members of this family may also be involved. Inactivation of TCF-7 (former TCF-1) leads to development of intestinal polyps . Expression of LEF1 is found in tumors but not in normal colon tissue . Each of these proteins can recognize the same TCF/LEF consensus binding site and, therefore, might bind alleles of rs6983267. High degree of similarity between TCF/LEF proteins may lead to cross-reactivity in chromatin immunoprecipitation (ChIP) assays. Thus, other TCF/LEF factors should also be examined for their effect on regulation of MYC expression.
In conclusion, SNP rs6983267 within the 8q24 region has been established as one of the strongest genetic risk factors for development of at least two types of cancer. Identification of functional mechanisms of this association is the highest priority of cancer genetics and would mean a significant step forward towards understanding of cancer pathogenesis and development of better diagnostic and therapeutic approaches. Our results provide new insights into the regulation of MYC expression by TCF7L2. However, further studies are needed to investigate alternative molecular mechanisms that can explain the association between rs6983267 and cancer risk.
We thank Yi-Ping Fu for help with statistical analysis. The study was supported by the intramural research program of NCI/NIH (LPO) and by an R21 award to JLH (1R21DK078029-01). We would like to thank Cullan Welch, Nikki Usher, and Marj Carlson for their technical assistance with the tissue processing.
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