The Biochemical Basis of The miR-21 Expression by The Mu-Opioid Receptor

Abstract

Opioid receptors are members of the superfamily of seven transmembrane G protein-coupled receptors (GPCRs) which share several structural and functional characteristics. There are 3 subtypes of opioid receptors, designated μ (MOR), δ (DOR), and κ (KOR) opioid receptors, have been found in the immune, nervous, gastrointestinal and other tissues. We have attempted to clarify the nature of MOR-induced signal transduction pathways in leukocytes. We found that the activation of MOR leads to a significant induction of ERK phosphorylation in peripheral blood mononuclear cells from normal donors using the MOR-selective agonist DAMGO. We are also interested in determining the role of this signaling pathway in the regulation of the immune response. Recent experiments using selective inhibitors suggests that the activation of ERK involves a pathway composed of Raf, Ras, and MEK1/2 kinases, but is independent of PI3 kinase. After treatment of multiple protein kinase inhibitors we found the PKC inhibitor Go-6983 and PLC inhibitor U73122 could also inhibit ERK phosphorylation in MOR stable line (HEK-MOR). According to the results from the Go-6983 and H-89 inhibitor treatment experiments, we found PKCμ/PKD1, a family member of Protein Kinase D, may be involved in MOR-induced ERK phosphorylation. We also found PKCμ/PKD1 S916 phosphorylation after MOR activation and the PKCμ specific inhibitor CID755673 inhibited the MOR-mediated ERK activation. ERK phosphorylation activated several transcription factors in human monocytes, the activation of transcription factors has been proved to induce miRNA expression. We have initiated a series of experiments to study the regulation of miRNA expression by MOR in human monocytes. We found miR-21, miR-155, miR-29a, miR-20b expression were significantly up-regulated following morphine treatment, and morphine-induced miR-21 expression is down-regulated following pretreatment with the ERK inhibitor U0126 and PKD inhibitor CID755673 in human primary monocytes. The results suggest that morphine-induced MOR activation results in up-regulation of miRNA expression human monocytes and this may regulate monocyte and/or macrophage function thought PKCμ/Ras/Raf/ERK signaling pathway.