Estrogen receptor alpha interaction with estrogen response element half-sites from the rat prolactin gene.
Estrogen regulation of the rat prolactin gene requires sequences within the DNase I hypersensitive site II (HSII). We have used overexpressed mouse estrogen receptor alpha (ERalpha) protein to study interactions of ERalpha with an imperfect estrogen response element (ERE) and four ERE half-site sequences from HSII. We confirmed that ERalpha has higher affinity for ERE half-sites than for the imperfect ERE. As expected, the imperfect ERE formed a complex with ERalpha similar to that between mERalpha and a consensus ERE in gel shift assays. The ERalpha complex with half-sites, however, had faster mobility on a 4% polyacrylamide gel than the ERalpha complex with a consensus ERE, indicating that the complexes had different compositions. Ferguson analysis revealed that the ERalpha/half-site complex had a larger molecular weight and higher negative charge than the ERalpha/consensus ERE complex. Similar results were observed with purified human ERalpha, showing that the ERalpha/half-site complex contained only ERalpha and oligonucleotides. These results are best explained by a model in which a dimer of ERalpha is bound to two half-site oligonucleotides. We propose that two ERalpha dimers may interact with the four ERE half-sites in HSII to influence estrogen regulation of this gene.
Anderson
I
I
Department of Biochemistry, University of Wisconsin, 433 Babcock Drive, Madison, Wisconsin 53706, USA.
Gorski
J
J
D000818:Animals; D001665:Binding Sites / Q000235:genetics; D004591:Electrophoresis, Polyacrylamide Gel; D004967:Estrogens / Q000235:genetics* / Q000378:metabolism; D005786:Gene Expression Regulation; D006801:Humans; D051379:Mice; D011388:Prolactin / Q000235:genetics*; D051381:Rats; D011960:Receptors, Estrogen / Q000096:biosynthesis / Q000378:metabolism*; D020218:Response Elements / Q000235:genetics*; D014819:Vitellogenins / Q000235:genetics; D014981:Xenopus
D004967:Estrogens; D011960:Receptors, Estrogen; D014819:Vitellogenins; D011388:Prolactin