Antigen cross-presentation at single-cell resolution

Antigen cross-presentation by dendritic cells (DCs) to CD8+ cytotoxic T cells (CTLs) is a key step in the immunological cascade. Although the overall sequence of events at DC synapses has been studied at the level of entire tissues, the molecular dynamics of cross-presentation, CTL activation and reprogramming at the level of individual cellular interactions remain incompletely understood. Studying direct physical interactions between cells presents greater challenges than analysing isolated individual cells separately. Previous approaches have relied on antibody-based methods, limiting the discovery of novel ligand-receptor interactions at the transcriptome-wide level. Here, we investigate the molecular and temporal changes characteristic of DC maturation and synapses in two steps: i) We first analyse the transcriptomes of single DCs over time as they uptake ovalbumin peptide, and discovered that foreign antigen uptake increases DC susceptibility to viral infection. ii) We then coculture these primed DCs with ovalbumin-specific OT1 CTLs and capture real-time transcriptomic profiles of single DCs, CTLs, and intentional DC-CTL doublets. Comparative analyses highlight the role of Rab and SNARE proteins in antigen cross-presentation. The methodology developed in this study provides a high-throughput, low-barrier approach to investigate the dynamic molecular basis in various cellular communication systems.