A Novel Signaling Pathway of Nitric Oxide on Transcriptional Regulation of Mouse κ Opioid Receptor Gene

Abstract

Nitric oxide (NO) suppressed the transcription of the mouse kappa opioid receptor (KOR) gene, mediated by a rapid downregulation of c-myc gene expression. KOR was constitutively expressed in postnatal day 19 (P19) embryonal carcinoma stem cells and is suppressed by NO donors [sodium nitroprusside (SNP), 3-morpholinosydnonimine-1, and S-nitrosoglutathione] in P19 stem cells within 4 hr. The suppression was reversed by 2-phenyl-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide, an NO scavenger, but could not be blocked by dithiothreitol, ruling out S-nitrosylation as the underlying mechanism. The suppressive effect of NO on KOR occurred at the level of gene transcription, mediated by E boxes located in promoters I and II of this gene. Protein-DNA complexes that formed on these E boxes contained c-myc; c-myc expression was suppressed by NO in P19 stem cells within 2 hr of treatment. Furthermore, chromatin immunoprecipitation demonstrated reduced c-myc binding to the E boxes and hypoacetylation of histone H3 on the chromatin of endogenous KOR promoters in P19 stem cells treated with SNP. It is proposed that NO regulates KOR at the level of gene transcription, mediated by a rapid suppression of c-myc gene expression and its binding to KOR promoters, and followed by chromatin hypoacetylation of and reduced transcription from KOR promoters in P19 stem cells. A novel pathway mediating the potential interplay between NO and opioid systems is discussed.

Netrin-1 Signaling RegulatesDe NovoProtein Synthesis of κ Opioid Receptor by Facilitating Polysomal Partition of Its mRNA

“…DRG neurons were prepared as described previously (Zheng et al, 2001;Stucky et al, 2002;Bi et al, 2003) and plated on 0.1 g/l poly-D-lysine-coated plates. P19 embryonal carcinoma cells were prepared as described previously (Park et al, 2002). Reporter assay was performed also as described previously (Hu et al, 2002).…”

“…Protein samples from P19 cells were prepared as described previously (Park et al, 2002). In brief, cell pellet was resuspended in immunoprecipitation buffer and sonicated.…”

“…Three opioid receptor types, , ␦, and , are known, and they belong to the super family of G-protein-coupled receptors (Minami and Satoh, 1995;Law et al, 1999;Pasternak, 2004). Regulation of their expression has been extensively examined, primarily at the level of gene transcription regulated by vitamin A, cytokines (Bi et al, 2001;Park et al, 2002;Wei and Loh, 2002;Hu et al, 2004), and specific transcription factors (Kraus et al, 2001;Kim et al, 2004). They also often involve chromatin remodeling events that occur over a period of time and are more permanent (Park et al, 2005).…”

Antiexudative Effects of Opioids and Expression of κ- and δ- Opioid Receptors during Intestinal Inflammation in Mice: Involvement of Nitric Oxide

“…Thus, we have recently demonstrated that nitric oxide derived from iNOS was implicated in the enhanced antitransit effects of morphine as well as in the enhanced transcription of MOR gene observed during intestinal inflammation . In contrast, nitric oxide suppressed the transcription of KOR gene in stem cells cultures (Park et al, 2002). No studies have been carried out to evaluate the effects of nitric oxide in the antiexudative effects of KOR and DOR agonists during intestinal inflammation.…”

Nuclear Factor κB Signaling in Opioid Functions and Receptor Gene Expression

“…No direct studies have been carried out to determine whether NF-κB signaling is involved in kor gene expression in immune cells. However, in the mouse embryonic carcinoma P19 cells, the NO donor sodium nitroprusside (SNP) was able to block c-Myc-mediated kor gene expression in retinoic acidtreated cells (Park et al 2002). Furthermore, nitration of NF-κB (Park et al 2005) is involved in the inactivation of c-myc gene expression in P19 cells (Park and Wei 2003), indicating that inhibition of NF-κB signaling could be the mechanism of SNP-mediated kor gene expression in retinoic acid-treated P19 cells (Law et al 2004;Park et al 2002).…”