Analyses of the genomic methylation status of the human cyclin A1 promoterby a novel real-time PCR-based methodology

The role of CpG methylation in the regulation of tissue-specific gene expre ssion is highly controversial. Cyclin A1 is a tissue-specifically expressed gene that is strongly methylated in non-expressing tumor cell lines. We ha ve established a novel real-time PCR method to quantitate genomic CpG methy lation of the cyclin A1 promoter. Genomic DNA samples from different human organs were treated with bisulfite and amplified with methylation-specific primers and with primers amplifying methylated as well as non-methylated DN A. PCR product quantitation was obtained by using a fluorogenic probe label ed with FAM and TAMRA. These analyses demonstrated that the human cyclin A1 promoter mas methylated in kidney, colon, spleen, testis, and small intest ine, but not in brain, liver, pancreas, or heart. Expression of cyclin A1 w as predominantly found in testis. Low level expression of cyclin A1 was pre sent in spleen, prostate, leukocytes, colon, and thymus. Taken together, ou r data provide evidence that CpG methylation patterns of the human cyclin A 1 promoter in human organs do not generally correlate with cyclin A1 gene e xpression in vivo. (C) 2001 Federation of European Biochemical Societies. P ublished by Elsevier Science B.V. All rights reserved.