Fig. 5.

Effect of PKCγ and of activated Gα subunits to segregate the HINT1-RGSZ2 complex. In vitro, PKCγ exhibits a moderate capacity to disrupt the interaction between RGSZ2 and HINT1 proteins. GST-RGSZ2 (100 nM) and HINT1 (200 nM) were incubated in the absence or presence of increasing concentrations of active PKCγ. RGSZ2 was then precipitated with glutathione sepharose (GS) and the associated HINT1 homodimer was evaluated (GS or GST did not precipitate HINT1, see Fig. 3b). In the presence of activated Gαi2/zGTPγS subunits, a sub-effective concentration of PKCγ (30 nM) produced segregation of the RGSZ2–HINT1 complex. The assay was carried out as above. In the lower panel, GαGTPγS subunits were used at 100 nM and the data are the mean ± SEM from two independent assays. *Significantly different from the corresponding control (C), ANOVA-Student–Newman–Keuls test; p < 0.05