The estrogen receptor alpha (ERalpha) is a ligand-dependent transcription factor that regulates a large number of genes in many different target tissues and is important in the development and progression of breast cancer. ERalpha-mediated transcription is a complex process regulated at many different levels. The interplay between ligand, receptor, DNA sequence, cofactors, chromatin context, and post-translational modifications culminates in transcriptional regulation by ERalpha. Recent technological advances have allowed the identification of ERalpha target genes on a genome-wide scale. In this review, we provide an overview of the progress made in our understanding of the different levels of regulation mediated by ERalpha. We discuss the recent advances in the identification of the ERalpha-binding sites and target gene network and their clinical applications.