Temporal mixture modelling of single-cell RNA-seq data resolves a CD4+ T cell fate bifurcation

Abstract Differentiation of naive CD4 + T cells into functionally distinct T helper subsets is crucial for the orchestration of immune responses. Due to multiple levels of heterogeneity and multiple overlapping transcriptional programs in differentiating T cell populations, this process has remained a challenge for systematic dissection in vivo . By using single-cell RNA transcriptomics and computational modelling of temporal mixtures, we reconstructed the developmental trajectories of Th1 and Tfh cell populations during Plasmodium infection in mice at single-cell resolution. These cell fates emerged from a common, highly proliferative and metabolically active precursor. Moreover, by tracking clonality from T cell receptor sequences, we infer that ancestors derived from the same naive CD4 + T cell can concurrently populate both Th1 and Tfh subsets. We further found that precursor T cells were coached towards a Th1 but not a Tfh fate by monocytes/macrophages. The integrated genomic and computational approach we describe is applicable for analysis of any cellular system characterized by differentiation towards multiple fates. One Sentence Summary Using single-cell RNA sequencing and a novel unsupervised computational approach, we resolve the developmental trajectories of two CD4 + T cell fates in vivo , and show that uncommitted T cells are externally influenced towards one fate by inflammatory monocytes.