Activator of G protein signalling I (AGS1) is a Ras-like protein that affects signalling through heterotrimeric G proteins. Previous in vitro studies suggest that AGS1 can bind to G(alpha)-GDP subunits and promote nucleotide exchange, leading to activation of intracellular signalling pathways. This model is consistent with in vivo evidence demonstrating that AGS1 activates both G(alpha)- and G(beta gamma)-dependent pathways in the absence of ligand. However, it does not easily explain how AGS1 blocks G(beta gamma)-dependent, but not G(alpha)-dependent, signalling following receptor activation. We have used yeast two hybrid analysis and co-immunoprecipitation studies in mammalian cells to demonstrate a direct interaction between AGS1 and the G(beta 1) subunit of heterotrimeric G proteins. The interaction is specific for G(beta 1), and involves the cationic region of AGS1 and the C-terminal region of G(beta 1). Possible implications of this novel interaction for the activity of AGS1 are discussed. (c) 2005 Elsevier Inc. All rights reserved.