title: Cholesterol inhibits the nuclear entry of estrogen receptor activation factor (E-RAF) and its dimerization with the nonactivated estrogen receptor (naER) in goat uterus.
abstract: An alternative form of estrogen receptor isolated from goat uterus, the nonactivated estrogen receptor (naER), has no DNA-binding function, although it is closely similar to the classical estrogen receptor (ER) in its hormone binding affinity and specificity. The naER dimerizes with a DNA binding protein, estrogen receptor activation factor (E-RAF). The heterodimer binds to the DNA. Assays carried out during the purification of E-RAF showed that an endogenous inhibitor that is heat stable and dialyzable bound to the E-RAF and prevented the formation of the heterodimer. The inhibitor has been isolated and purified. GC-MS analysis identifies this molecule to be cholesterol. Circular dichroism measurement has shown that the high-affinity binding of cholesterol to E-RAF results in subtle changes in the secondary and the tertiary structure of the protein. The E-RAF with altered conformation fails to dimerize with the naER. Instead of facilitating E-RAF entry into the nucleus, dimerization with the naER prevents it. Similarly, cholesterol binding blocks the nuclear entry of the protein, showing that E-RAF with altered conformation is incapable of interaction with the nuclear pore complex/membrane proteins. The naER-E-RAF heterodimer remains at the nuclear periphery, incapable of further transport. These results indicate the possibility that the dimerization between naER and the E-RAF takes place only within the nuclear compartment. The observation that cholesterol binding prevents nuclear entry of the E-RAF reflects the similarity of E-RAF with the sterol regulatory element (SRE) binding protein that enters the nucleus and binds to SRE only when the intracellular level of cholesterol remains low.
authors.affiliation: Rajiv Gandhi Centre for Biotechnology, Jagathy, Thiruvananthapuram 695 014, Kerala, India. rgcbt@md2.vsnl.net.in
authors.forename: R V
authors.initials: RV
authors.lastname: Thampan
authors.forename: A
authors.initials: A
authors.lastname: Zafar
authors.forename: N S
authors.initials: NS
authors.lastname: Imam
authors.forename: S
authors.initials: S
authors.lastname: Sreeja
authors.forename: K
authors.initials: K
authors.lastname: Suma
authors.forename: M
authors.initials: M
authors.lastname: Vairamani
mesh_terms: D000818:Animals; D002467:Cell Nucleus / Q000378:metabolism*; D002499:Centrifugation, Density Gradient; D002783:Cholestenones / Q000138:chemical synthesis; D002784:Cholesterol / Q000494:pharmacology*; D002855:Chromatography, Thin Layer; D002942:Circular Dichroism; D019281:Dimerization; D005260:Female; D016650:Fluorescein-5-isothiocyanate / Q000378:metabolism; D008401:Gas Chromatography-Mass Spectrometry; D006041:Goats; D013058:Mass Spectrometry; D011485:Protein Binding; D011506:Proteins / Q000037:antagonists & inhibitors / Q000302:isolation & purification / Q000378:metabolism*; D011960:Receptors, Estrogen / Q000302:isolation & purification / Q000378:metabolism*; D014599:Uterus / Q000378:metabolism*
chemical_list: D002783:Cholestenones; D011506:Proteins; D011960:Receptors, Estrogen; C030311:4,6-cholestadien-3-one; D002784:Cholesterol; D016650:Fluorescein-5-isothiocyanate