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Figure 1 | Molecular Cancer

Figure 1

From: Promoter hypermethylation of the SFRP2 gene is a high-frequent alteration and tumor-specific epigenetic marker in human breast cancer

Figure 1

SFRP2 expression and promoter methylation in breast cancer cell lines. (A) SFRP2 mRNA expression in benign and malignant cell lines was determined by RT-PCR. All but one malignant cell line (Hs578T) completely lacked SFRP2 mRNA expression. Of the two benign breast cell lines (MCF10A and HMEC) only HMEC cells were found to express abundant SFRP2 mRNA. In a commercially available human normal breast tissue cDNA SFRP2 expression was clearly detectable, while expression was strongly reduced in a corresponding cDNA of human malignant breast tissue. (B) Genomic structure of the human SFRP2 gene on chromosome 4q31. Bioinformatic analysis revealed three CGIs (light blue), located within the SFRP2 promoter (left CGI), the 5' untranslated region (UTR; central CGI) and the coding sequence (CDS; right CGI). Methylation of the central CGI was explored by MSP. Circles indicate CpG sites; filled circles represent MSP forward (MSP-F) and reverse (MSP-R) primer hybridization sites. Indicated positions are related to the transcription start site (+1) initiating the 5'-UTR. (C) MSP was performed with bisulfite-treated DNA from the same breast cell lines as in A. DNA bands in lanes labeled with U indicate PCR products amplified with primers recognizing unmethylated SFRP2 promoter sequence. DNA bands in lanes labeled with M represent amplification products with methylation-specific primers. Five out of eight mammary tumor cell lines exhibit complete promoter methylation (MDA-MB-231, MCF7, MDA-MB-453, BT20 and BT474), two cell lines show partial SFRP2 methylation (SKBR3 and T47D). In Hs578T, only unmethylated SFRP2 promoter sequence could be detected, like it was also found in benign HMEC cells. In addition, lymphocyte DNA from a healthy donor did not reveal SFRP2 methylation. GAPDH served as cDNA loading control; NTC represents the no template control.

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