Specific methylation events contribute to the transcriptional repression of the mouse tissue inhibitor of metalloproteinases-3 gene in neoplastic cells

The tissue inhibitor of metalloproteinases-3 (TIMP-3) gene is specifically down-regulated in neoplastic cells of the mouse JB6 progression model, sugg esting a role for TIMP-3 inactivation in neoplastic progression. On the bas is of 5-azacytidine reversal, the mechanism for this down-regulation appear s to involve changes in the methylation state of the TIMP-3 promoter. Altho ugh total genomic methylation levels are comparable, specific differences i n the methylation of the TIMP-3 promoter were observed between preneoplasti c and neoplastic JB6 cells at three HpaII sites, with preneoplastic cells b eing less methylated. Expression of antisense methyltransferase in a neopla stic JB6 variant known to be hypermethylated in TIMP-3 resulted in reactiva tion of the endogenous TIMP-3 gene and restoration of hypomethylated status to the three implicated HpaII sites. Thus, hypermethylation at specific se quences in the TIMP-3 promoter appears to contribute to the silencing of th e gene in neoplastic cells.