Memory CD8+ T cells mediate early pathogen-specific protection through localized delivery of antigen-dependent chemokines and IFNgamma to clusters of inflammatory monocytes

While cognate antigen-driven clonal expansion is a cardinal feature of memory CD8+ T cell effective response, inflammatory cytokines also orchestrates their rapid activation and production of IFNᵧ, contributing to innate mechanisms of protection. Despite antigen-independent secretion, IFNᵧ accounts for a large part of antigen-dependent protection in a multitude of infections, yet how cognate antigen enables memory CD8+ T cell-derived IFNᵧ to achieve host protection remains unknown. Herein, using distinct models of immunization, we establish that cognate antigen (Ag) recognition by memory CD8+ T cells on conventional CD11chi dendritic cells initiates their coordinated production of a burst of chemokines (CCL3, CCL4, XCL1) under the transcriptional control of IRF4. Using intravital microscopy imaging, we further reveal that memory CD8+ T cells undergo Ag-mediated arrest in splenic red pulp clusters of CCR2+ monocytes to deliver them with IFNᵧ- and Ag-dependent chemokine-potentiating microbicidal activities. These results establish that effective memory CD8+ T cell responses require coordinated stepwise processes that quickly restrict pathogen growth and optimize the local delivery of effector molecules before clonal expansion occurs. We adoptively transferred 2,000 naive OT-I (ova-specific) and 50,000 gBT-I (HSV2-specific) CD8+ T cells to mice subsequently immunized with 10,000 Listeria monocytogenes expressing the Ova and the gB antigen (Lm-Ova-gB). 6 weeks later, mice were challenged or not with 100,000 Lm-Ova. Spleens were harvested 8 hrs post challenge and OT-I and gBT-I memory cells were enriched by negative selection (dynabeads) and 1,000 OT-I and 1,000 gBT-I were flow-sorted (Aria III) into lysis buffer before proceeding to first strand cDNA synthesis and amplification, and library (Illumina) preparation for sequencing (RNA-seq).