FIG. 6.
The cancer risk-associated rs6983267(G) allele is linked to enhanced c-MYC expression. (A) Diagram of the linked polymorphisms at c-MYC SNPs rs6983267 and rs4645953 in DLD-1 cells as determined by 3C analysis. (B) Allele-specific c-MYC expression as evaluated by PCR analysis of the c-MYC rs4645953 SNP in DLD-1 cells. Genomic DNA exhibits heterozygosity at the rs4645953 SNP (DLD-1 DNA profile). RT-PCR detects twofold-enhanced expression of the c-MYC rs4645953(C) allele (DLD-1 RNA profile). RT-PCR detects the same twofold-enhanced expression of the c-MYC rs4645953(C) allele after knockdown of beta-catenin expression with siRNA (DLD-1 + β-catenin siRNA profile). Sequencing profiles accurately detect allele-specific expression because transient transfection with cloned full-length c-MYC genes carrying the different rs4645953 alleles (C and T alleles) gives a 1:1 ratio of expression levels when the c-MYC genes are mixed in a 1:1 ratio (rs4645953 C/T 1:1 profile). Mixing the c-MYC rs4645953 C and T alleles in a 2:1 ratio gives a corresponding 2:1 ratio of expression levels as determined by RT-PCR (rs4645953 C/T 2:1 profile). (C) RT-PCR quantitation of c-MYC allele-specific expression in DLD-1 cells before and after knockdown of beta-catenin (β-cat) expression by using siRNA. (D) Knockdown of beta-catenin expression gives a corresponding reduction in c-MYC expression. GAPDH, glyceraldehyde-3-phosphate dehydrogenase.