Specification of the direction of adhesive signaling by the integrin β cytoplasmic domain

Abstract

Integrin adhesion receptors can signal in two directions: first, they can regulate cellular behaviors by modulating cellular signaling enzymes ("outside-in signaling"); second, cells can regulate the affinity of integrins ("inside-out signaling") by such pathways. Integrin β cytoplasmic domains (tails) mediate both types of signaling, and Src family kinases (SFKs) and talin, which bind to β tails, are important for integrin signaling. Here, we utilized "homology scanning" mutagenesis to identify β tail mutants selectively defective in c-Src binding and found that amino acid exchanges affecting a combination of an Arg and Thr residue in the integrin β3 tail control the binding specificity for SFKs but have no effect on talin binding. Using β tail mutants at these residues, we found that SFK binding to integrin β tails is dispensable for inside-out signaling but is obligatory for cell spreading, a marker of outside-in signaling. Conversely, we found that point mutations that disrupt talin binding abolish integrin activation, but they do not inhibit SFK binding to the β3 tail or the initiation of outside-in signaling once the integrins are in a high affinity form. Thus, we show that inside-out and outside-in integrin signaling are mediated by distinct and separable interactions of the integrin β tails. Furthermore, based on our results, it is possible to discern the relative contributions of the direction of integrin signaling on biological functions in cell culture and, ultimately, in vivo. © 2005 by The American Society for Biochemistry and Molecular Biology, Inc.