Metastasis tumor antigen 1 (MTA1), a novel candidate metastasis-associated gene, is known to increase the migration and invasion of various tumor cells in vitro. It also plays an important role in tumorigenesis and tumor aggressiveness of breast cancer. Estrogen receptor alpha (ERα) plays an important role in the etiology of breast cancer and has been widely accepted as a prognostic marker for breast cancer and a response predictor for endocrine therapy. The ERα gene methylation has been linked to the lack of ERα expression in breast cancer. The aim of the study is to assess the correlation between the ERα methylation and MTA1 expression in breast cancer and further to investigate whether the repressed ERα methylation can downregulate the expression of MTA1 in vitro. In general, we found ERα methylation had significant correlation with the MTA1 expression (p < 0.05) in female patients of breast cancer (n = 102) by methylation-specific polymerase chain reaction and immunohistochemistry. To gain a deeper insight into the molecular mechanism underlying the relation between MTA1 and ERα methylation, we treated the invasive breast cancer cell lines with the demethylating agent, found the downregulation of MTA1 protein expression, and mRNA with the unmethylation of ERα (p < 0.05). And the invasive ability of breast cancer cells was significantly positively associated with MTA1 expression. These unique findings have greatly extended our current knowledge about the relation between ERα methylation and MTA1 expression. These data strongly support the hypothesis that methylation is involved in the relation between MTA1 and ERα in breast cancer.