The estrogen receptor is one of the most important transcription factors in breast tumor growth and development. We, and others, have previously shown that hypoxia induces rapid ER alpha protein degradation by a proteasome-mediated pathway in breast cancer cells, which is linked with ER alpha activation. However, no report has shown the effect of hypoxia on ESR1 gene regulation at the transcriptional level. In this report, we show that hypoxia repressed the expression of ER alpha mRNA in MCF-7 and T47D human breast cancer cells, but not in human endometrial Ishikawa cells, although ERa degradation under hypoxia was also observed in Ishikawa cells. This indicates that ESR1 transcriptional repression and ER alpha protein downregulation by hypoxia are regulated by distinct mechanisms. Repression of ESR1 gene transcription occurred at the transcriptional level as a result of decreased recruitment of RNA polymerase II at the proximal promoter of the ESR1 locus in response to stabilization of the HIF-1 alpha protein under hypoxia. Our data show that hypoxia induces repression of the ESR1 gene, which may facilitate hormone insensitivity in the tumor microenvironment. (C) 2011 Elsevier Inc. All rights reserved.