Inhibition of Talin-induced Integrin Activation by a Double-hit Stapled Peptide

Abstract

Integrins are ubiquitously expressed cell-adhesion proteins. Talin is required for integrin activation through an inside-out signaling pathway, during which talin is recruited to the plasma membrane (PM) by RAP1 directly or through its effector, RAP1-Interacting Adaptor Molecule (RIAM). RIAM also activates talin from autoinhibition by binding to talin head domain. A helical talin-binding segment (TBS) in RIAM mediates both talin activation and recruitment by binding to two distinct sites in talin head and rod domains respectively. The bi-specificity of the TBS fragment allows us to develop a new strategy to suppress talin-induced integrin activation through a "double-hit" approach. We designed an experimental peptidomimetic inhibitor by engineering a hydrocarbon "staple" in the helical TBS fragment to mask the integrin binding site in the talin head. The stapled peptide (S-TBS) exhibits a stronger binding affinity with talin and inhibits talin:integrin interaction. Crystal structure of S-TBS in complex with talin rod domains reveals an interface that overlaps with the TBS-binding site in talin. Consistently, S-TBS also exhibits an inhibitory effect on TBS:talin-rod interaction during the PM recruitment of talin. Importantly, S-TBS possesses excellent cell permeability and inhibits integrin activation in a talin-dependent manner. Hence, our results present a novel approach to design a new class of intracellular inhibitors targeting integrin functions.