Agonist-induced Desensitization of the μ Opioid Receptor Is Determined by Threonine 394 Preceded by Acidic Amino Acids in the COOH-terminal Tail

Background: Neuropeptide FF 2 receptors interact with -opioid receptors and decrease their activity. Results: The phosphorylation pattern of MOP receptor is similar after homologous (DAMGO) and heterologous (neuropeptide FF) stimulation. Conclusion: GPCR kinase 2 (GRK2) contributes to the NPFF-induced phosphorylation and loss of function of MOP receptor. Significance: GRK2-mediated transphosphorylation within receptor heteromers could play a role in heterologous desensitization of GPCRs.

Section: Discussioncontrasting

confidence: 56%

GRK2 Protein-mediated Transphosphorylation Contributes to Loss of Function of μ-Opioid Receptors Induced by Neuropeptide FF (NPFF2) Receptors

Moulédous

Froment

Dauvillier

et al. 2012

“…More specifically, segments of the carboxyl tail and specific residues contained therein have been identified for both (20,23,24) and ␦ (25) receptors as important for agonist-induced internalization. We hypothesized therefore, that the inability of the ␦ receptor to internalize when co-expressed with the receptor indicated that the relevant region of the carboxyl tail that mediated ␦ opioid receptor internalization was somehow occluded.…”

Section: Discussionmentioning

confidence: 99%

A Role for the Distal Carboxyl Tails in Generating the Novel Pharmacology and G Protein Activation Profile of μ and δ Opioid Receptor Hetero-oligomers

Fan

Varghese

Nguyen

et al. 2005

Self Cite

Opioid receptor pharmacology in vivo has predicted a greater number of receptor subtypes than explained by the profiles of the three cloned opioid receptors, and the functional dependence of the receptors on each other shown in gene-deleted animal models remains unexplained. One mechanism for such findings is the generation of novel signaling complexes by receptor hetero-oligomerization, which we previously showed results in significantly different pharmacology for and ␦ receptor hetero-oligomers compared with the individual receptors. In the present study, we show that deltorphin-II is a fully functional agonist of the -␦ heteromer, which induced desensitization and inhibited adenylyl cyclase through a pertussis toxin-insensitive G protein. Activation of the -␦ receptor heteromer resulted in preferential activation of G␣ z , illustrated by incorporation of GTP␥ 35 S, whereas activation of the individually expressed and ␦ receptors preferentially activated G␣ i . The unique pharmacology of the -␦ heteromer was dependent on the reciprocal involvement of the distal carboxyl tails of both receptors, so that truncation of the distal receptor carboxyl tail modified the ␦-selective ligand-binding pocket, and truncation of the ␦ receptor distal carboxyl tail modified the -selective binding pocket. The distal carboxyl tails of both receptors also had a significant role in receptor interaction, as evidenced by the reduced ability to co-immunoprecipitate when the carboxyl tails were truncated. The interaction between and ␦ receptors occurred constitutively when the receptors were co-expressed, but did not occur when receptor expression was temporally separated, indicating that the hetero-oligomers were generated by a co-translational mechanism.The endogenous opioid systems mediate important physiological functions related to pain perception, locomotion, motivation, reward, autonomic function, immune modulation, and hormone secretion. Opioid receptors, mediating the actions of the opioid peptides, belong to the family of G protein-coupled receptors (GPCRs) 2 and have distinct pharmacological profiles with discrete but overlapping distributions in brain (reviewed Refs. 1 and 2). The pharmacology of the opioid receptors obtained in brain and other tissues has consistently predicted a greater number of receptor subtypes such as 1, 2, and ␦1, ␦2, which are not explained by the individual pharmacological profiles obtained for the three cloned opioid receptors, , ␦, and , when expressed individually in heterologous systems. One possible mechanism for these findings, gaining increasing credence, has been that of direct receptorreceptor interactions creating novel signaling units and generating distinct post-receptor functional effects.The ability of GPCRs to form homo-oligomers and hetero-oligomers has been described by us and others (reviewed in Refs. 3-5). We have provided evidence for the direct interaction of and ␦ opioid receptors to form hetero-oligomers, with generation of novel pharmacology and G protein coupling properti...

“…Thus, several Ser/Thr residues within the COOH terminus of opioid receptors have been suggested to be involved in the regulation of these receptors. Thr 394 , Ser 353 , and Ser 369 have been shown to be important for the regulation of -opioid receptors (20,34,35), ␦-opioid receptors (18,19), and -opioid receptors (36), respectively, although phosphorylation of these residues has not been determined. Recently, a T394A mutation was shown to significantly block DAMGO-induced MOR phosphorylation to approximately 10% of the wild type receptor and impair receptor desensitization in Chinese hamster ovary cells (37), indicating that this residue is a phosphorylation site.…”

Section: Discussionmentioning

confidence: 99%

“…Several kinases have been postulated to be involved in opioid receptor phosphorylation including protein kinase A (38,39), protein kinase C (33,40), GRKs (25,34), mitogen-activated protein kinase (41,42), calmodulin kinase II (43,44), and tyrosine kinase (32 (21). Although MOR and DOR have differences in their C-tail primary sequence, it is interesting to note that a common motif encompassing agonistinduced phosphorylated sites exists for both receptor subtypes.…”

Section: Discussionmentioning

confidence: 99%

Phosphorylation of Ser363, Thr370, and Ser375 Residues within the Carboxyl Tail Differentially Regulates μ-Opioid Receptor Internalization

Kouhen

Burd

Erickson-Herbrandson

et al. 2001

117

126

Prolonged activation of opioid receptors leads to their phosphorylation, desensitization, internalization, and down-regulation. To elucidate the relationship between -opioid receptor (MOR) phosphorylation and the regulation of receptor activity, a series of receptor mutants was constructed in which the 12 Ser/Thr residues of the COOH-terminal portion of the receptor were substituted to Ala, either individually or in combination. All these mutant constructs were stably expressed in human embryonic kidney 293 cells and exhibited similar expression levels and ligand binding properties. to Ala accelerated MOR internalization kinetics. The present data show that the basal phosphorylation of MOR could play a role in modulating agonist-induced receptor internalization kinetics. Furthermore, even though -receptors and ␦-opioid receptors have the same motif encompassing agonist-induced phosphorylation sites, the different agonist-induced internalization properties controlled by these sites suggest differential cellular regulation of these two receptor subtypes.Opioid alkaloids, as well as endogenous opioid peptides, exert their multiple biological effects on target tissues by interacting with specific cell surface receptors including the ␦-, -, and -opioid receptors (1). These opioid receptors belong to the superfamily of G protein-coupled receptors (GPCRs).1 The -opioid receptor (MOR) serves as the principle physiological target for most clinically important opioid analgesics, such as morphine and fentanyl (2, 3). Although many opioid alkaloids exert their pharmacological effects via MOR, their binding affinity for the receptor and potency to activate the receptor do not always correspond to their abilities to induce tolerance (4 -8). This suggests that other cellular processes that modulate MOR responsiveness, such as receptor desensitization and internalization, may contribute to opioid tolerance and dependence. Like many other GPCRs, opioid receptors are regulated by agonist-dependent processes and undergo receptor phosphorylation, desensitization, internalization, and down-regulation (1). Interestingly, in addition to the subtype-specific regulation of opioid receptors (9 -12), individual opioid receptors are differentially regulated by distinct opioid agonists (5-8, 13, 14). In the case of MOR, opioid agonists demonstrating equivalent ability to activate receptor signaling exhibit remarkable differences in their abilities to functionally desensitize (5, 6) and induce internalization of the receptor in both transfected cells and neurons (7,8,(13)(14)(15). However, the detailed molecular events underlying this differential regulation of MOR by distinct agonists remain unclear. Using chimeric, truncated, or mutated opioid receptors, several studies reported the crucial role of the C-tail of opioid receptors in regulating their activities and trafficking (11, 16 -20). Whereas several potential phosphorylation sites were suggested to be involved in regulating the activity and trafficking of opioid receptors, the actual ...