Rapid Nitric Oxide-Mediated S-Nitrosylation of Estrogen Receptor: Regulation of Estrogen-Dependent Gene Transcription

Nitric oxide (NO) and estrogen receptor (ER) are both important mediators of signal transduction in cardiovascular and reproductive tissues. In this study, we evaluated NO-mediated S-nitrosylation of ER and assessed the effect of this structural modification on transcription-related functions of ER. We have found selective inhibitory effects of NO on specific binding of ER to specific estrogen-responsive elements (ERE) that can be reversed in the presence of the reducing agent, DTT, thus suggesting that S-nitrosylation of thiolate-zinc centers may occur within the ER molecule. Furthermore, we examined inhibitory effects of NO on ER-dependent transcriptional activity by using an ERE-driven reporter gene system. By monitoring biophysical changes in the structure of NO-treated or untreated human recombinant ERα, we obtained evidence for the formation of S-nitrosothiols in the ER molecule. In addition, we have detected specific S-nitrosylation of cysteine residues within the ER molecule by immunodetection of S-nitrosocysteine moieties in ER. Collectively, these findings suggest an important physiological role for NO in modification of human ER structure by S-nitrosylation, an effect that leads, in turn, to impaired DNA-binding activity of ER and subsequent blockade of estrogen-dependent gene transcription. Thus, NO-induced S-nitrosylation of ER can occur at cysteine residues that coordinate Zn2+within the two major DNA-binding Zn-finger domains of ER, resulting in selective inhibition of DNA-binding at specific ERE. This cross-communication between NO and ER may favor activation of rapid (nongenomic) signaling pathways and subsequent modulation of downstream genomic activity.