Specificity of Gβγ signaling to Kir3 channels depends on the helical domain of pertussis toxin-sensitive Gα subunits

Abstract

Acetylcholine signaling through muscarinic type 2 receptors activates atrial G protein-gated inwardly rectifying K+ (Kir3) channels via the βγ subunits of G proteins (Gβγ). Different combinations of recombinant Gβγ subunits have been shown to activate Kir3 channels in a similar manner. In native systems, however, only Gβγ subunits associated with the pertussis toxin-sensitive Gαi/o subunits signal to K+ channels. Additionally, in vitro binding experiments supported the notion that the C terminus of Kir3 channels interacts preferentially with Gαi over Gαq. In this study we confirmed in two heterologous expression systems a preference of Gαi over Gαq in the activation of K + currents. To identify determinants of Gγβ signaling specificity, we first exchanged domains of Gαi and Gαq subunits responsible for receptor coupling selectivity and swapped their receptor coupling partners. Our results established that the G proteins, regardless of the receptor type to which they coupled, conferred specificity to Kir3 activation. We next tested signaling through chimeras between the Gαi and Gαq subunits in which the N terminus, the helical, or the GTPase domains of the Gα subunits were exchanged. Our results revealed that the helical domain of Gαi (residues 63-175) in the background of Gαq could support Kir3 activation, whereas the reverse chimera could not. Moreover, the helical domain of the Gαi subunit conferred "Gαi- like" binding of the Kir3 C terminus to the Gαq subunits that contained it. These results implicate the helical domain of Gαi proteins as a critical determinant of Gγβ signaling specificity. © 2007 by The American Society for Biochemistry and Molecular Biology, Inc.