Agonist-Mediated Tyrosine Phosphorylation of Isoforms of the Shc Adapter Protein by the δ Opioid Receptor

Maximally effective concentrations of the opioid agonist D-ala 2-D-leu 5-enkephalin resulted in some 2–3-fold enhancement of tyrosine phosphorylation of the p52 Shc adapter protein in a clone of Rat-1 fibroblasts transfected to express stably the murine δ opioid receptor. More limited modifications of the tyrosine phosphorylation status of the p46 and p66 forms of Shc were observed in parallel. Epidermal growth factor caused some 10–12-fold enhancement of tyrosine phosphorylation of p52 Shc and marked increases in the p46 and p66 forms. The effect of D-ala 2-D-leu 5-enkephalin was prevented by pretreatment of the cells with pertussis toxin and was not observed in non-transfected parental fibroblasts whereas the effect of epidermal growth factor was still manifest in both these situations. Half-maximal effects of D-ala 2-D-leu 5-enkephalin on p52 Shc tyrosine phosphorylation were produced with sub-nanomolar concentrations, in agreement with previous results on the tyrosine phosphorylation of p44 MAPK (Burt et al., 1996). p52 Shc became tyrosine phosphorylated more rapidly than p44 MAPK in response to D-ala 2-D-leu 5-enkephalin and its enhanced tyrosine phosphorylation was maintained for at least 10 min.