Estrogen receptor alpha (ERalpha) interacts with basal transcription factors, coregulatory proteins, and chromatin modifiers to initiate transcription of the target genes. We have identified a novel interaction between ERalpha and the DNA repair protein 3-methyladenine DNA glycosylase (MPG) thereby providing a functional link between gene expression and DNA repair. Interestingly, the ERalpha-MPG interaction was enhanced by the presence of estrogen response element (ERE)-containing DNA. In vitro pull-down assays indicated that the interaction of ERalpha with MPG was direct and occurred through the DNA- and ligand-binding domains and the hinge region of the receptor. More importantly, endogenously expressed ERalpha and MPG from MCF-7 cells coimmunoprecipitated with ERalpha- and MPG-specific antibodies. The ERalpha-MPG interaction had functional consequences on the activities of both proteins. ERalpha increased MPG acetylation, stabilized the binding of MPG with hypoxanthine-containing oligos, and enhanced MPG-catalyzed removal of hypoxanthine from DNA. In turn, MPG dramatically stabilized the interaction of ERalpha with ERE-containing oligos, decreased p300-mediated acetylation of the receptor, and reduced transcription of simple and complex ERE-containing reporter plasmids in a dose-dependent manner. Our studies suggest that recruitment of MPG to ERE-containing genes influences transcription and plays a role in maintaining integrity of the genome by recruiting DNA repair proteins to actively transcribing DNA.