Structural basis for T cell regulation by the CTLA-4/B7-2 complex

Abstract

CTLA-4, like CD28 and ICOS, is a preformed dimeric receptor and, as such, does not have available to it the classical signaling mechanism of ligand-induced dimerization. We reasoned that the transmission of inhibitory signals by CTLA-4 would most likely rely on either a (1) B7-induced conformational change in individual CTLA-4 homodimers that would be transmitted to its cytoplasmic tails to recruit downstream signaling molecules or (2) higher-order clustering of multiple CTLA-4 and B7 molecules that would trigger CTLA-4 signaling cascades in the responding T cell. In order to elucidate the signaling mechanism utilized by CTLA-4, we developed a bacterial expression system for the production of recombinant CTLA-4 and B7-2 molecules for biochemical and crystallographic studies. We report the structures of the disulfide-linked homodimer of human CTLA-4 and the complex formed between CTLA-4 and the receptor-binding domain of human B7-2.;The structure of the CTLA-4B7-2 complex reveals that the CTLA-4 dimer does not undergo any significant conformational changes upon B7-2 binding, ruling this is out as a potential signaling mechanism. In contrast to previous studies that showed that B7-1 is dimeric in solution and crystalline form, these studies demonstrate that B7-2 is strictly monomeric in solution. However, in the crystal, B7-2 forms a similar dimer to that observed in the unliganded B7-1 structure, which suggests that B7-2 undergoes receptor-induced dimerization. Like CTLA-4, the B7-2 dimer observed in the crystal places its ligand binding sites distal to the dimer interface, providing the potential for each B7-2 dimer to bind two independent CTLA-4 molecules. The bivalent potential of both CTLA-4 and B7-2 is realized by the formation of a periodic assembly of dimeric molecules, characterized by a 103 A repeat, that extends throughout the entirety of the crystal. Direct observation of this CTLA-4B7-2 network provides a model for the periodic organization of these molecules within the immunological synapse and suggests a distinct mechanism for signaling by dimeric cell surface receptors. Given the similarities in their B7 binding sites and their modes of dimerization, most aspects of the organized assembly observed for the CTLA-4/B7 complex should be operative for both CD28/B7 and ICOS/B7h complexes, potentially establishing the signaling mechanism paradigm for this costimulatory receptor-ligand superfamily. (Abstract shortened by UMI.).