Figure 4. Extending the D2 C-terminus also inverts the gating regulation.

(a) D2 = K0-25 is inhibited by 5 μM dopamine (Dopa). D2=K+9M2-25 contains an extended D2 C-terminus with the last 9 residues from M2 and is activated by 5 μM Dopamine. The first transmembrane domain (TMD0) of SUR1 is co-expressed for boosting the surface expression of the ICCR. (b) Basal currents of dopaminergic ICCRs show lack of surface expression of D2=K+9M2-25. Co-expression with the N-terminal transmembrane domain of SUR1 (TMD0) restores the surface expression. Extended C-terminus of D2 with 9 alanines is noted D2=K+9Ala-25 and with 9 C-terminal residues from the human β2 adrenergic receptor is noted D2=K+9β2-25. *: compared to Non-Injected P < 0.02; ns (not significant) P = 6.10⁻². (c) Average in % of current amplitude ± s.e.m. induced by 5 μM dopamine. *: P < 10⁻³ (ref.: D2 + K∆). (d) Alignment of the C-terminal sequences of the human receptors M2 (blue), D2 (green) and β2 adrenergic (brown) receptors. The position of the helix VIII is shown above the sequences and the terminal cysteines are numbered and depicted as palmitoylated. The long C-terminus of the β2 adrenergic receptor is partially represented. The 3 different extensions of the D2 C-terminus are shown in the lower panel. (e) TEVC recording of D2=K+9M2-25 + Kir3.4* showing D2-mediated activation of Kir3.4* channel by 5 μM dopamine (green arrow) and the antagonism (red arrow) by 5 μM sulpiride. (f ) Average current change induced by 5 μM dopamine (green bar) and 5 μM dopamine + 5 μM sulpiride (red bar) on D2=K+9M2-25 + Kir3.4*. * P = 9.10⁻³ between the two bars. n = 6.