CD2 expression acts as a quantitative checkpoint for immunological synapse structure and T-cell activation

Abstract

The CD2 receptor has been described as an adhesion and costimulatory receptor on T cells. Here, transcriptional profiling of colorectal cancers (CRC) revealed a negative correlation between CD2 expression and "exhausted CD8 + Tcells" gene signatures. Furthermore, we detected reduced surface CD2 levels in exhausted CD127 low PD-1 hi CD3 + CD8 + tumour infiltrating lymphocytes (TILs) in CRC.We describe a CD2 expression-level-dependent switch in CD2-CD58 localization between central and peripheral domains in the immunological synapse (IS). A peripheral "CD2 corolla" formed when CD2 surface expression was sufficiently high and its cytoplasmic domain intact. The corolla recruited other ligated receptors like CD28, boosted recruitment of activated Src-family kinases (pSrc), LAT and PLC-γ in the IS and consequently T-cell activation in response to a tumour antigen. Corolla formation and pSrc in the IS increased linearly with CD2 expression, whereas pSrc signals were reduced by high, "exhausted-like" levels of PD-1, which invaded the corolla. These results suggest two levels of inhibition of Src-family kinases in CD3 + CD8 + TILs: reduced CD2 expression and high PD-1 expression.

In silico characterization of mechanisms positioning costimulatory and checkpoint complexes in immune synapses

“…Integrin-mediated adhesion is an active process regulated by interactions with the F-actin transport network (6), whereas the sIGSF members operate through highly multivalent microdomains and are less dependent on cellular energy (7). When a live T cell forms an interface with a planar supported lipid bilayer (SLB) containing freely mobile ICAM-1, CD58 and pMHC, the individual complexes are organized into a layered, radially symmetrical pattern with central TCR-pMHC cluster (synaptic cleft and central supramolecular activation cluster, cSMAC), surrounded by an integrin adhesion ring (peripheral or pSMAC), and fringed by sIGSF microdomains in the distal SMAC (dSMAC), resembling flower petals known as corolla (5,(8)(9)(10)(11). cSMAC and pSMAC formation driven by F-actin mediated centripetal transport (10,12,13) and intermittent coupling of TCR-pMHC to the F-actin network (14) with a constant force (15).…”

“…cSMAC and pSMAC formation driven by F-actin mediated centripetal transport (10,12,13) and intermittent coupling of TCR-pMHC to the F-actin network (14) with a constant force (15). The corolla pattern is completely distinct from that formed by costimulatory and checkpoint complexes, like CD28-CD80 and PD1-PDL1, which also occupy TCR microclusters and the outer annulus of the cSMAC (3,11,(16)(17)(18). We have previously shown that cSMAC localization of both TCR-pMHC and costimulatory complexes can be simulated when engaged TCR and CD28 interact independently with the Factin transport network (19).…”

“…We have previously shown that cSMAC localization of both TCR-pMHC and costimulatory complexes can be simulated when engaged TCR and CD28 interact independently with the Factin transport network (19). Strikingly, addition of CD2-CD58 complexes to this system leads to corolla formation and relocation of the costimulatory complexes to the corolla in the dSMAC (11). Thus, it is important to understand the forces that regulate corolla formation.…”

“…CD2 interactions with hCD58 or mCD48 are low affinity with fast dissociation kinetics (23)(24)(25)(26). The long, unstructured and highly conserved cytoplasmic domain of CD2 is rich in polyproline motifs (PP) that contribute to Src family kinase activation (10,11,(27)(28)(29)(30). Similar unstructured domains with multiple PP in combination with proteins containing multiple SH3 domains can undergo concentration dependent liquid-liquid phase separation (31).…”

“…Nevertheless, it plays an important role for TCR signaling (34), positively regulating T cell activation via dephosphorylation of the inhibitory C-terminal tyrosine of p56lck and p59fyn proteins (35)(36)(37)(38), which also interact with CD2 (27). Importantly, CD45 molecules are excluded from the TCR-pMHC microclusters during the initial moments of IS formation and eventually completely relocate to the outer region of the IS, the dSMAC (35,39,40), where the corolla forms (11). This suggests that CD45 may play a role in the organization of the corolla/dSMAC, although no previous studies have addressed this question to the best of our knowledge.…”