The endogenous antigen-specific CD8+ T cell repertoire is composed of unbiased and biased clonotypes with differential fate commitment.

Generating balanced populations of CD8+ effector and memory T cells is necessary for immediate and durable immunity to infections and cancer. Yet, a definitive understanding of how a diverse CD8+ T cell repertoire differentiates remains unclear. We identified several hundred TCR clonotypes that constitute the polyclonal response against a single antigen and found that a majority of TCR clonotypes were highly biased towards memory or effector fate. TCR-intrinsic biases were not stochastic and were dominant over environmental cues. Differential gene expression analysis of memory- or effector-biased TCR clonotypes showed bifurcation of differential fates at the early effector stage. Additionally, phylogenetic analysis revealed that memory-biased clonotypes retain their fate preferences in subclonal populations but effector-biased subclones can switch to memory fate. Our study highlights that the polyclonal CD8+ T cell response is a composite of unbiased and biased clonotypes with varying capacity to incorporate environmental cues in their cell fate decisions. Overall design: scRNA-seq, scRAGE-seq (scTCR-seq) and scCARLIN barcode sequencing were performed on splenic CD8+ T cells isolated from CARLIN mice infected with VSV-OVA acute viral infection. scRNA-seq for transcriptional profiling of the cells and the scCARLIN barcode sequencing to uncover the edits in the CARLIN array were performed using Illumina sequencing. scTCR-seq was performed using Nanopore technology as done in the original RAGE-seq paper (https://www.nature.com/articles/s41467-019-11049-4).