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Updated: May 9, 2026
Systems Biology of Metabolic Regulation by Estrogen Receptor Signaling in Breast Cancer
Published on: March 17, 2016
Characterization of estrogen response element binding proteins as biomarkers of breast cancer behavior
Traci L Kruer1, Timothy D Cummins, David W Powell
1Hormone Receptor Laboratory, Department of Biochemistry & Molecular Biology, Brown Cancer Center, University of Louisville, Louisville, KY 40292, USA; Institute for Molecular Diversity & Drug Design, University of Louisville, Louisville, KY 40292, USA.
Background:
While investigating estrogen response element (ERE) binding properties of human estrogen receptor-α (hERα) in breast cancer cytosols, other ERE-binding proteins (ERE-BP) were observed.
Design And Methods:
Recognition properties of ERE-BP were evaluated by electrophoretic mobility shift assays (EMSA) with ERE sequences of the 5'-flanking region of the estrogen responsive gene vitellogenin A2 (VitA2). Cytosols were incubated 16 h, 4 °C with [32P]ERE sequences and separated by EMSA. A method of estimating ERE-BP levels was developed by measuring band intensity from EMSA profiles, expressed in digital light units (DLU)/μg protein and normalized to total DLU. ERE-BP were purified by affinity chromatography and EMSA, and then identified by mass spectrometry.
Results:
ERE-BP in cytosols did not supershift in the presence of anti-hERα or anti-hERβ antibodies recognizing different ER epitopes suggesting that they are not fragments of either receptor isoform. ERE-BP competed with hERα for binding to VitA2-ERE. Increased levels of ERE-BP DNA-binding activities measured in 310 cytosols prepared from breast cancer biopsies correlated with decreased patient survival. Strikingly, breast cancer patients with ER negative status and high ERE-BP expression exhibited the poorest disease-free and overall survival. After purification, ERE-BP were identified as Ku70 (XRCC6) and Ku80 (XRCC5) using mass spectrometry. ERE-BP were confirmed to be Ku70/80 by supershift assay.
Conclusion:
Presence of these novel ERE-binding proteins in a breast carcinoma is a strong predictor of poor prognosis. Our results suggest that ERE-BP, identified as Ku70/Ku80, in cytosols prepared from breast carcinoma biopsies are useful biomarkers for assessing risk of breast cancer recurrence.
