Transmembrane helices of integrin α and β subunits have been implicated in the regulation of integrin activity. Two mutations, glycine-708 to asparagine-708 (G708N)and methionine-701 to asparagine-701, in the transmembrane helix of the β3 subunit enabled integrin αIIbβ3 to constitutively bind soluble fibrinogen. Further characterization of the G708N mutant revealed that it induced αIIbβ3 clustering and constitutive phosphorylation of focal adhesion kinase. This mutation also enhanced the tendency of the transmembrane helix to form homotrimers. These results suggest that homomeric associations involving transmembrane domains provide a driving force for integrin activation. They also suggest a structural basis for the coincidence of integrin activation and clustering.