The process of p15 CpG island methylation induced by granulocyte-macrophage colony-stimulating factor (GM-CSF) was investigated, using MO7e cells. The cells proliferating in response to GM-CSF+fetal bovine serum (FBS) were almost fully methylated in the p15 CpG island. The withdrawal of both GM-CSF and FBS for 48 h reduced the cell viability, and increased the frequency of alleles with completely or partially demethylated CpG sites by approximately 50%. Viable cells were responsible for this epigenetic change. The add-back of GM-CSF restored the methylation. Seventy-two hours withdrawal of GM-CSF+FBS followed by 24-h exposure to inhibitors for DNA methyltransferase (DNMT) and histone deacetylase (HDAC) caused the demethylation of nearly all CpG sites in the p15 CpG island on every allele sequenced. When GM-CSF was re-added after 96-h treatment, the cells exhibited p15 transcriptional silencing via the methylation. The initial methylation event encompassed the entire CpG island. No new methylated alleles appeared in the coexistence of the DNMT and HDAC inhibitors. Taken together, GM-CSF may be able to induce de novo methylation of the p15 gene, using HDAC(s) as well as DNMT(s).