B and T lymphocyte attenuator (BTLA) provides an inhibitory signal to B and T cells. Previously, indirect observations suggested that B7x was a ligand for BTLA. Here we show that BTLA does not bind B7x; instead, we identify herpesvirus entry mediator (HVEM) as the unique BTLA ligand. BTLA bound the most membrane-distal cysteine-rich domain of HVEM, distinct from regions where the ligands LIGHT and lymphotoxin-alpha bound HVEM. HVEM induced BTLA tyrosine phosphorylation and association of the tyrosine phosphatase SHP-2 and repressed antigen-driven T cell proliferation, providing an example of reverse signaling to a non-tumor necrosis factor family ligand. The conservation of the BTLA-HVEM interaction between mouse and human suggests that this system is an important pathway regulating lymphocyte activation and/or homeostasis in the immune response.

cytokines ͉ immune evasion ͉ T lymphocytes ͉ costimulation E fficient activation and differentiation of T cells depends on recognition of antigen and cooperating signals (cosignaling) that provoke either positive or inhibitory effects. Inhibitory pathways help maintain immune tolerance to self tissues. In the absence of inhibitory signals or with sustained positive cosignaling, tolerance can be overridden, leading to autoimmune responses. Two major groups of cosignaling receptors are recognized, those with an Ig-like fold, such as CTLA-4 (1), CD28 (2), PD1 (3), and B and T lymphocyte attenuator (BTLA) (4, 5), and those belonging to the TNF receptor (TNFR) superfamily, including OX40, 41BB, CD27, CD30, and herpesvirus entry mediator (HVEM; TNFR superfamily 14) among others (6-9). Generally, positive cosignaling receptors in the Ig family act by sustaining antigen receptor-associated kinase activity, whereas their inhibitory counterparts contain an immunoreceptor tyrosine-based inhibitory motif that recruits phosphatases [e.g., Src homology 2 (SH2) domain phosphatase-1; SH2 domaincontaining inositol polyphosphate 5-phosphatase] attenuating antigen receptor signaling (1, 2, 10). By contrast, the cosignaling TNFRs activate serine kinases promoting expression of survival and proinflammatory genes through the transcription factors NF-B and activator protein 1, whereas some other TNFRs induce apoptosis, negatively regulating T cells by cellular elimination (6).A unique inhibitory cosignaling pathway for T cells was recently defined (11,12), which involves the engagement of BTLA by HVEM, connecting the Ig and TNFR cosignaling families. HVEM binding activates tyrosine phosphorylation of the immunoreceptor tyrosine-based inhibitory motif in BTLA and induces the association with the protein tyrosine phosphatases Src homology domain 1 and 2 required for inhibitory signaling (13). However, HVEM can also act as a positive cosignaling receptor (reviewed in ref. 8) by binding TNF-related ligands LIGHT (TNF superfamily 14) and lymphotoxin ␣ (LT␣, TNF superfamily 2) (14). A fourth ligand of HVEM is envelope glycoprotein D (gD) of herpes simplex virus (HSV; ␣-herpesvirus) from which its name was derived (15, 16).Thus, HVEM has the potential to serve as a molecular switch mediating either positive or inhibitory signaling, depending on which of its four ligands are bound. Moreover, the directionality of signaling is not clear, as the hierarchy of ligand occupancy and relative affinities of the interactors are not well defined. The N-terminal extracellular region of HVEM is composed of four pseudorepeats of a cysteine-rich domain (CRD), characteristic of the TNFR superfamily; each repeat contains three disulfide bonds that fold into complex loops depending in part on the spacing of the cysteines (17). Mutagenesis studies (18) . Domain-swapping experiments revealed the CRD1 of HVEM was sufficient to mediate BTLA binding (11). These observations raised the issue of the location of the BTLA binding site relative to gD and LIGHT.Here, w...