METHYLATED CPG POINTS IDENTIFIED WITHIN MAGE-1 PROMOTER ARE INVOLVED IN GENE REPRESSION

The MAGE-1 gene, expressed in some tumors of different histological or igins, codes far a tumor antigen recognized by cytotoxic T lymphocytes . The gene is not expressed in normal tissues with the exception of te stes. The present study was designed to investigate the relationship b etween methylation of the MAGE-1 promoter and inactivation of the MAGE -1 gene. We examined the extent to which MAGE-1 B'B promoter sequences are methylated in tumor-cell lines, in order to determine whether met hylation correlates with MAGE-1 expression. Using methylation-sensitiv e restriction analysis followed by polymerase chain reaction (PCR), we found an inverse correlation between methylation of the IMAGE-1 B'B r egion and MAGE-1 expression. An unmethylated state was identified in D NA from sperm and some tumor-cell lines of different origins. In contr ast, a hypermethylation state was found in leukocytes and other MAGE-1 non-expressing cells. Furthermore, treatment with 5-aza-2'-deoxycytid ine, a demethylating agent, induced MAGE-1 expression in tumor-cell li nes in which we found no direct relation between transcriptional activ ity of the B'B region and MAGE-1 expression. Binding of the nuclear fa ctors to the B'-methylated probe was strongly inhibited, indicating th at methylation of cytosine interferes directly in the binding of trans criptional factors. (C) 1996 Wiley-Liss, Inc.