The gene of mouse kappa opioid receptor (KOR) utilizes two promoters, P1 and P2. P1 is active in various brain areas and constitutively in P19 mouse embryonal carcinoma cells. P2 is active in limited brain stem areas of adult animals and only in late differentiated cells of P19 induced for neuronal differentiation in the presence of nerve growth factor (NGF). NGF response of P2 was found to be mediated by a specific binding site for transcription factor activation protein 2 (AP2) located in P2. Electrophoretic gel shift assay showed specific binding of this AP2 site by AP2β, but not AP2α. Knockdown of endogenous AP2β with siRNA abolished the stimulating effect of NGF on the expression of transcripts driven by P2. Binding of endogenous AP2β on the endogenous KOR P2 chromatin region was also confirmed by chromatin immunoprecipitation. The effect of NGF was inhibited by LY2942002 (phosphatidylinositol 3-kinase, PI3K inhibitor), suggesting that PI3K was involved in signaling pathway mediating the effect of NGF stimulation on KOR P2. The chromatin of P2 in P19 was found to be specifically modified following NGF stimulation, which included demethylation at Lys9 and dimethylation at Lys4 of histone H3 and was consistent with the increased recruitment of RNA polymerase II to this promoter. This study presents the first evidence for epigenetic changes occurred on a specific KOR promoter triggered by NGF in cells undergoing neuronal differentiation. This epigenetic change is mediated by recruited AP2β to this promoter and involves the PI3K system.
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|Published - Feb 19 2008
Bibliographical noteFunding Information:
This work was supported partially by NIH grants DA11190, DA11806, DK54733, DK60521 and K02-DA13926 to L.-N.W., DA000564, DA001583, DA011806, and K05-DA070554 to H.H.L. and by a seed grant of DA011806 to S.W.P.
- P19 neuronal differentiation
- kappa opioid receptor
- retinoic acid