Activator of G protein signaling 3 is a guanine dissociation inhibitor for Gαi subunits

Activator of G protein signaling 3 (AGS3) is a newly identified protein shown to act at the level of the G protein itself. AGS3 belongs to the GoLoco family of proteins, sharing the 19-aa GoLoco motif that is a G alpha (i/o) binding motif. AGS3 interacts only with members of the G alpha (i/o) subfamily. By surface plasmon resonance, we found that AGS3 binds exclusively to the GDP-bound form of G alpha (i3). In GTP gammaS binding assays, AGS3 behaves as a guanine dissociation inhibitor (GDI), inhibiting the rate of exchange of GDP for GTP by G alpha (i3). AGS3 interacts with both G alpha (i3) and G alpha (o) subunits, but has GDI activity only an G alpha (i3), not on G alpha (o),. The fourth GoLoco motif of AGS3 is a major contributor to this activity. AGS3 stabilizes G alpha (i3) in its GDP-bound form, as it inhibits the increase in tryptophan fluorescence of the G alpha (i3)-GDP subunit stimulated by AlF4-. AGS3 is widely expressed as it is detected by immunoblotting in brain, testis, liver, kidney, heart, pancreas, and in PC-12 cells. Several different sizes of the protein are detected. By Northern blotting. AGS3 shows 2.3-kb and 3.5-kb mRNAs in heart and brain, respectively. suggesting tissue-specific alternative splicing. Taken together, our results demonstrate that AGS3 is a GDI. To the best of our knowledge, no other GDI has been described for heterotrimeric G proteins. Inhibition of the G alpha subunit and stimulation of heterotrimeric G protein signaling, presumably by stimulating G beta gamma, extend the possibilities for modulating signal transduction through heterotrimeric G proteins.