DAPLE protein inhibits nucleotide exchange on Gα_s and Gα_q via the same motif that activates Gαi

Besides being regulated by G-protein– coupled receptors, the activity of heterotrimeric G proteins is modulated by many cytoplasmic proteins. GIV/Girdin and DAPLE (Dvl-associating protein with a high frequency of leucine) are the best-characterized members of a group of cytoplasmic regulators that contain a Gα-binding and -activating (GBA) motif and whose dysregulation underlies human diseases, including cancer and birth defects. GBA motif–containing proteins were originally reported to modulate G proteins by binding Gα subunits of the G_i/o family (Gα_i) over other families (such as G_s, G_q/11, or G_12/13), and promoting nucleotide exchange in vitro. However, some evidence suggests that this is not always the case, as phosphorylation of the GBA motif of GIV promotes its binding to Gα_s and inhibits nucleotide exchange. The G-protein specificity of DAPLE and how it might affect nucleotide exchange on G proteins besides Gα_i remain to be investigated. Here, we show that DAPLE’s GBA motif, in addition to Gα_i, binds efficiently to members of the G_s and G_q/11 families (Gα_s and Gα_q, respectively), but not of the G_12/13 family (Gα₁₂) in the absence of post-translational phosphorylation. We pinpointed Met-1669 as the residue in the GBA motif of DAPLE that diverges from that in GIV and enables better binding to Gα_s and Gα_q. Unlike the nucleotide-exchange acceleration observed for Gα_i, DAPLE inhibited nucleotide exchange on Gα_s and Gα_q. These findings indicate that GBA motifs have versatility in their G-protein–modulating effect, i.e. they can bind to Gα subunits of different classes and either stimulate or inhibit nucleotide exchange depending on the G-protein subtype.