Novel DNA motif binding activity observed in vivo with an estrogen receptor α mutant mouse.
Estrogen receptor α (ERα) interacts with DNA directly or indirectly via other transcription factors, referred to as "tethering." Evidence for tethering is based on in vitro studies and a widely used "KIKO" mouse model containing mutations that prevent direct estrogen response element DNA- binding. KIKO mice are infertile, due in part to the inability of estradiol (E2) to induce uterine epithelial proliferation. To elucidate the molecular events that prevent KIKO uterine growth, regulation of the pro-proliferative E2 target gene Klf4 and of Klf15, a progesterone (P4) target gene that opposes the pro-proliferative activity of KLF4, was evaluated. Klf4 induction was impaired in KIKO uteri; however, Klf15 was induced by E2 rather than by P4. Whole uterine chromatin immunoprecipitation-sequencing revealed enrichment of KIKO ERα binding to hormone response elements (HREs) motifs. KIKO binding to HRE motifs was verified using reporter gene and DNA-binding assays. Because the KIKO ERα has HRE DNA-binding activity, we evaluated the "EAAE" ERα, which has more severe DNA-binding domain mutations, and demonstrated a lack of estrogen response element or HRE reporter gene induction or DNA-binding. The EAAE mouse has an ERα null-like phenotype, with impaired uterine growth and transcriptional activity. Our findings demonstrate that the KIKO mouse model, which has been used by numerous investigators, cannot be used to establish biological functions for ERα tethering, because KIKO ERα effectively stimulates transcription using HRE motifs. The EAAE-ERα DNA-binding domain mutant mouse demonstrates that ERα DNA-binding is crucial for biological and transcriptional processes in reproductive tissues and that ERα tethering may not contribute to estrogen responsiveness in vivo.
Receptor Biology (S.C.H., W.W., K.J.H., B.P., K.S.K.), Laboratory of Reproductive and Developmental Toxicology, Biostatistics Branch (L.L.), Integrative Bioinformatics (S.A.G., D.F.), Laboratory of Structural Biology (L.C.P.), National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, North Carolina 27709; Department of Molecular and Cellular Biology (C.A.R., R.B.L., F.J.D.), Baylor College of Medicine, Houston, Texas 77030; and Department of Molecular Biology of the Cell (G.S.), German Cancer Research Center, 69121 Heidelberg, Germany.
Sylvia C
SC
Hewitt
Leping
L
Li
Sara A
SA
Grimm
Wipawee
W
Winuthayanon
Katherine J
KJ
Hamilton
Brianna
B
Pockette
Cory A
CA
Rubel
Lars C
LC
Pedersen
David
D
Fargo
Rainer B
RB
Lanz
Francesco J
FJ
DeMayo
Günther
G
Schütz
Kenneth S
KS
Korach
D000818:Animals; D001483:Base Sequence; D016384:Consensus Sequence; D004958:Estradiol / Q000502:physiology; D047628:Estrogen Receptor alpha / Q000235:genetics* / Q000378:metabolism; D005260:Female; D000090062:Kruppel-Like Factor 4; D051741:Kruppel-Like Transcription Factors / Q000235:genetics / Q000378:metabolism; D057507:Mice, 129 Strain; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D020125:Mutation, Missense; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism; D010641:Phenotype; D011485:Protein Binding; D020218:Response Elements; D015533:Transcriptional Activation; D014599:Uterus / Q000378:metabolism
D047628:Estrogen Receptor alpha; C414172:KLF15 protein, human; C000715467:KLF4 protein, human; C000715468:Klf4 protein, mouse; D000090062:Kruppel-Like Factor 4; D051741:Kruppel-Like Transcription Factors; D009687:Nuclear Proteins; D004958:Estradiol