To study the role of the N-terminal region of the estrogen receptor (ER) in transcription activation and in DNA binding, we constructed a mutant of the Xenopus laevis ER which lacks amino acids 1-159 (XER160/586). In transient transfections, XER160/586 exhibited <10% of the activity of wild-type XER on a synthetic promoter containing two estrogen response elements (EREs). To examine transcriptional synergism by XER and by XER160/586, we determined the activity of promoters containing EREs and binding sites for either the vitellogenin activator, NF1, or API upstream activator protein. For the three promoters transcription by XER was 2.8-fold greater than expected for additive activities, and transcription by XER160/586 was 6.2-fold greater. These data demonstrate that an upstream activator protein bound near the promoter can partially compensate for the loss of the internal N-terminal (AF1) transactivation domain in XER160/586. Using a promoter interference assay to study the intracellular interaction between ER and the estrogen response element, we found that XER160/586 exhibited a significant increase in affinity for the ERE. Its low basal activity and enhanced affinity for the ERE make XER160/586 an effective dominant negative mutant. When co-expressed with wild-type XER at 1:1 and 5:1 ratios, XER160/586 suppressed the activity of wild-type XER by 57% and >80%, respectively.
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