title: Delta opioid activation of the mitogen-activated protein kinase cascade does not require transphosphorylation of receptor tyrosine kinases.
abstract: BACKGROUND
  In this study, we investigated the mechanism(s) by which delta opioids induce their potent activation of extracellular signal-regulated protein kinases (ERKs) in different cell lines expressing the cloned delta-opioid receptor (delta-OR). While it has been known for some time that OR stimulation leads to the phosphorylation of both ERK isoforms, the exact progression of events has remained elusive.
  
  
  RESULTS
  Our results indicate that the transphosphorylation of an endogenous epidermal growth factor receptor (EGFR) in the human embryonic kidney (HEK-293) cell line does not occur when co-expressed delta-ORs are stimulated by the delta-opioid agonist, D-Ser-Leu-enkephalin-Thr (DSLET). Moreover, neither pre-incubation of cultures with the selective EGFR antagonist, AG1478, nor down-regulation of the EGFR to a point where EGF could no longer activate ERKs had an inhibitory effect on ERK activation by DSLET. These results appear to rule out any structural or catalytic role for the EGFR in the delta-opioid-mediated MAPK cascade. To confirm these results, we used C6 glioma cells, a cell line devoid of the EGFR. In delta-OR-expressing C6 glioma cells, opioids produce a robust phosphorylation of ERK 1 and 2, whereas EGF has no stimulatory effect. Furthermore, antagonists to the RTKs that are endogenously expressed in C6 glioma cells (insulin receptor (IR) and platelet-derived growth factor receptor (PDGFR)) were unable to reduce opioid-mediated ERK activation.
  
  
  CONCLUSIONS
  Taken together, these data suggest that the transactivation of resident RTKs does not appear to be required for OR-mediated ERK phosphorylation and that the tyrosine-phosphorylated delta-OR, itself, is likely to act as its own signalling scaffold.
authors.affiliation: Department of Biological Science, Fordham University, Larkin Hall-Room 160, 440 East Fordham Road, Bronx, NY 14440, USA. hakramer@fordham.edu
authors.forename: H Kenneth
authors.initials: HK
authors.lastname: Kramer
authors.forename: Irma
authors.initials: I
authors.lastname: Onoprishvili
authors.forename: Matthew L
authors.initials: ML
authors.lastname: Andria
authors.forename: Kayane
authors.initials: K
authors.lastname: Hanna
authors.forename: Karina
authors.initials: K
authors.lastname: Sheinkman
authors.forename: Lisa B
authors.initials: LB
authors.lastname: Haddad
authors.forename: Eric J
authors.initials: EJ
authors.lastname: Simon
mesh_terms: D002460:Cell Line; D004743:Enkephalin, Leucine / Q000031:analogs & derivatives* / Q000494:pharmacology*; D004789:Enzyme Activation; D004815:Epidermal Growth Factor / Q000494:pharmacology; D066246:ErbB Receptors / Q000378:metabolism; D006801:Humans; D020935:MAP Kinase Signaling System / Q000187:drug effects*; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D009294:Narcotics / Q000494:pharmacology*; D010766:Phosphorylation; D020794:Receptor Protein-Tyrosine Kinases / Q000378:metabolism*; D011972:Receptor, Insulin / Q000378:metabolism; D017468:Receptors, Fibroblast Growth Factor / Q000378:metabolism; D017465:Receptors, Opioid, delta / Q000378:metabolism*; D017479:Receptors, Platelet-Derived Growth Factor / Q000378:metabolism
chemical_list: D009294:Narcotics; D017468:Receptors, Fibroblast Growth Factor; D017465:Receptors, Opioid, delta; D004743:Enkephalin, Leucine; D004815:Epidermal Growth Factor; C034318:enkephalin, Ser(2), Leu(5), Thr(6)-; D066246:ErbB Receptors; D020794:Receptor Protein-Tyrosine Kinases; D011972:Receptor, Insulin; D017479:Receptors, Platelet-Derived Growth Factor; D020928:Mitogen-Activated Protein Kinases