The conserved membrane-proximal region of an integrin cytoplasmic domain specifies ligand binding affinity

Abstract

Integrin affinities for ligands can change markedly via a process termed inside-out signaling. We expressed several truncations of the β3 cytoplasmic domain in conjunction with an 'activating' α subunit chimera, α(IIb)α(6B). Deletion of the 4 C-terminal residues of the β3 tail blocked inside-out signaling as assessed by the binding of an activation-specific antibody, PAC1. Several additional truncations remained in the low affinity state, but complete truncation (β3Δ717) caused PAC1 binding. Activation by this truncation mutant did not depend on the a subunit cytoplasmic domain and was resistant to inhibitors of cellular metabolism and the over-expression of an isolated β3 cytoplasmic domain. Since deletion of β3(Leu717- Asp723) results in a constitutively activated integrin, this membrane- proximal seven amino acids of the β3 cytoplasmic domain is required to maintain in a default low affinity state. The amino acid sequence of this region is conserved among integrins. Moreover, the conserved membrane- proximal sequence in α subunit tails seems to serve a similar function. Consequently, the conserved membrane-proximal regions of both integrin cytoplasmic domains control the ligand binding affinity of the extracellular domain.