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Nucleic Acids Research, Vol 27, Issue 20 4040-4049, Copyright © 1999 by Oxford University Press

ARTICLES

AP2alpha and AP2gamma: a comparison of binding site specificity and trans-activation of the estrogen receptor promoter and single site promoter constructs

LA McPherson and RJ Weigel
Department of Surgery, Stanford University, MSLS, Room P214, Stanford, CA 94305-5408, USA.

The AP2 transcription factors exhibit a high degree of homology in the DNA binding and dimerization domains. In this study, we methodically compared the binding specificity of AP2alpha and AP2gamma using PCR- assisted binding site selection and competitive gel shift assay and determined that the consensus binding site for both factors is(G)/(C)CCNN(A/)C(/G)(G)/(A)G(G/)C(/T.)The use of single site promoter constructs with either a high or low affinity site demonstrated a direct relationship between site affinity and transcriptional activation. Overexpression of AP2alpha and AP2gamma resulted in the activation of a low affinity binding site construct to levels comparable to those seen with a high affinity site construct at lower amounts of protein expression. Both AP2alpha and AP2gamma were able to trans-activate the cloned human estrogen receptor alpha promoter in ER- negative MDA-MB-231 cells through high affinity AP2 sites in the untranslated leader sequence. This provides a functional mechanism to explain the correlation between AP2 activity and estrogen receptor expression in breast cancer. Since there is overexpression of AP2 factors in breast cancer compared to normal breast epithelium, our results suggest that increased factor expression may activate a set of target genes containing lower affinity binding sites that would normally not be expressed in normal breast epithelium.
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