Activation of estrogen receptor-alpha by nitrite
Creator
Storchan, Geoffrey
Advisor
Martin, Mary Beth
Abstract
Recent studies show that the anion nitrite binds to and activates estrogen receptor-alpha (ERα). Mutational analysis and molecular modeling identified three potential nitrite binding sites in the ligand binding domain (LBD) of the receptor. Site A is formed by lys529 on helix H11 and asn532 in the loop between helices H11 and H12; site B is formed by his516 on helix H10 and lys520 on helix H11; and site C is formed by cys381 on helix H4 and his547 on helix H12. To determine the mechanism by which nitrite activates ERα, wild-type and mutant forms of ERα were tested for their ability to dissociate from the heat shock protein 90 (hsp90) complex, bind DNA, interact with coactivator, and recruit SRC-1 and RNA polymerase II. Results presented here demonstrate that, upon treatment with 1 μM nitrite, wild-type ERα dissociated from hsp90, was recruited to DNA, bound coactivator, recruited SRC-1 and RNA polymerase II to DNA and increased the expression of estrogen regulated genes. Site A mutants, K529A and N532A, did not dissociate from the hsp90-receptor complex, bind to DNA or bind coactivator. The site B mutants, H516A and K520A, dissociated from hsp90. However, H516A did not bind to DNA or recruit coactivator, while K520A bound to DNA, but did not recruit coactivator. The site C mutants, C381A and H547A, dissociated from hsp90. C381A failed to bind to DNA, while H547A was recruited to DNA but failed to bind coactivator. The results suggest a model whereby the interaction of nitrite with site A results in a conformational change at the interface of helix H11 and loop 11-12 that is necessary for the dissociation of hsp90. Site B is involved in the formation of a continuous helix between helices H10 and H11 that is necessary for binding to DNA. Site C, formed by amino acids located on helices H4 and H12, is involved in the recruitment to DNA as well as the formation of the coactivator binding site. Together, these findings represent a novel role for the anion nitrite in the activation of ERα.
Description
Ph.D.
Permanent Link
http://hdl.handle.net/10822/557516Date Published
2012Subject
Type
Publisher
Georgetown University
Extent
136 leaves
Related items
Showing items related by title, author, creator and subject.
-
Role of Calcium in Activation of Estrogen Receptor-Alpha
Divekar, Shailaja Dilip (Georgetown University, 2008)ABSTRACT Growth factors are thought to activate estrogen receptor-alpha ERα by phosphorylating the serines in the N-terminus of the receptor. However, this mechanism does not account for the conformational changes that ...