CTLA-4-dependent trogocytosis promotes the interactions between Tregs and antigen-presenting cells

Abstract

Regulatory T cells (Tregs) are responsible for the maintenance of immune homeostasis. They express the coinhibitory receptor CTLA-4 in high quantities, whose loss causes severe systemic autoinflammation in both mice and humans. CTLA-4 limits the expression of costimulatory CD80/CD86 proteins by antigen-presenting cells (APCs). However, the details of this mechanism are not fully understood. When we analyzed in vitro cocultures of CD80- or CD86-GFP expressing murine dendritic cells (DCs) with ex vivo sorted Tregs, we found that mutant Tregs, whose CTLA-4 lacked the cytoplasmic tail necessary for its endocytosis function, could still uptake CD80- or CD86-GFP from DCs via CTLA-4-dependent trogocytosis. Moreover, the specific CD80/86 uptake by CTLA-4, also facilitated the indirect transfer of membrane lipid particles and other surface proteins. Both flow cytometry and confocal microscopy revealed that immune synapse formation between Tregs and APCs was enhanced depending on CTLA-4 expression by Tregs. Confocal microscopy also demonstrated that uptaken CD80/CD86-GFP proteins colocalized with LFA-1 capping, an indicator of stable immune synapses. Furthermore, in vivo delivery of CTLA-4 WT and knockout (KO) Tregs into CD45.1+RAG2 KO mice showed that transfer of congenic CD45.1 protein from host cells to donor Tregs was significantly diminished in CTLA-4 KO Tregs compared to WT Tregs. The tail portion of CTLA-4 was also found to be dispensable for CD80/86 downregulation and suppressive functions of Tregs. In conclusion, CTLA-4-promoted immune synapse formation provides a stable platform between Tregs and APCs, which results in both depletion of CD80/CD86 via trogocytosis and indirect uptake of other surface proteins from APCs.