Role for G protein-coupled receptor kinase in agonist-specific regulation of mu-opioid receptor responsiveness.

The G protein-coupled mu-opioid receptor (mu OR) mediates the physiological effects of endogenous opioid peptides as well as the structurally distinct opioid alkaloids morphine and etorphine. An intriguing feature of mu OR signaling is the differential receptor trafficking and desensitization properties following activation by distinct agonists, which have been proposed as possible mechanisms related to opioid tolerance. Here we report that the ability of distinct opioid agonists to differentially regulate mu OR internalization and desensitization is related to their ability to promote G protein-coupled receptor kinase (GRK)-dependent phosphorylation of the mu OR. Although both etorphine and morphine effectively activate the mu OR, only etorphine elicits robust mu OR phosphorylation followed by plasma membrane translocation of beta-arrestin and dynamin-dependent receptor internalization. In contrast, corresponding to its inability to cause mu OR internalization, morphine is unable to either elicit mu OR phosphorylation or stimulate beta-arrestin translocation. However, upon the overexpression of GRK2, morphine gains the capacity to induce mu OR phosphorylation, accompanied by the rescue of beta-arrestin translocation and receptor sequestration. Moreover, overexpression of GRK2 also leads to an attenuation of morphine-mediated inhibition of adenylyl cyclase. These findings point to the existence of marked differences in the ability of different opioid agonists to promote mu OR phosphorylation by GRK. These differences may provide the molecular basis underlying the different analgesic properties of opioid agonists and contribute to the distinct ability of various opioids to induce drug tolerance.

Signal transduction pathways involved in the expression of the uridine diphosphoglucose pyrophosphorylase gene of Dictyostelium discoideum.

The uridine diphosphoglucose pyrophosphorylase (UDPGP1) gene of Dictyostelium discoideum is an excellent marker to study the pathways that control the expression of genes during development. We have previously shown that the UDPGP1 gene is regulated by exogenous cAMP acting on cell-surface cAMP receptors. Various steps in the signal transduction pathway between receptor stimulation and the induction of the gene can now be studied. Induction does not require the synthesis of intracellular cAMP, but does require new protein synthesis. By deletion and transformation with altered genes, two cis-acting sequences that are required for UDPGP1 expression have been identified. A GC-rich palindromic sequence located between -410 and -374 is essential for induction of the gene by extracellular cAMP, but not for its basal expression. A sequence element located between -374 and -337 is required for any basal expression of this gene. When the polarity of the palindromic sequence was reversed such that it resembled the H2K enhancer element, the gene could still be induced by exogenous cAMP. Two DNA binding activities were detected in gel mobility shift assays using a fragment containing both of the regulatory sequence elements of UDPGP1 gene. Transformation with a vector that resulted in the synthesis of anti-sense UDPGP1 RNA led to almost total elimination of the enzyme antigen and no detectable enzyme activity. However, these transformants developed normally, indicating that either UDPGP is not required for development or residual synthesis of UDPGP may be sufficient for normal development.