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Divergent clonal differentiation trajectories of T cell exhaustion

Daniel B. et al. (BioRxiv) DOI: 10.1101/2021.12.16.472900

Divergent clonal differentiation trajectories of T cell exhaustion


  • T cell exhaustion

  • Single cell sequencing

  • Chronic viral infection

Main Findings

To better characterize the heterogeneity of exhausted T (Tex) cell states, their clonal relationships, and molecular programming underlying their exhaustion, Daniel, B. et al. generated a single-cell multi-omic atlas of T cell exhaustion during chronic lymphocytic choriomeningitis virus (LCMV) infection. Through this high-dimensional profiling, they discovered a novel early effector and a novel late-stage killer cell lectin-like receptor (KLR) expressing Tex cell state. The early effector phenotype is epigenetically primed for T cell exhaustion, and the late-stage KLR Tex phenotype is concurrent with the terminal Tex phenotype.

By T cell clone tracing using paired single-cell RNA and TCR sequencing, they found that individual clones either preferentially acquired the terminal or KLR exhausted T cell phenotypes or developed into both phenotypes. They termed these T cell clone behaviors: terminal Tex biased, KLR Tex biased, and “divergent”, respectively. They further investigated the role of TCR affinity to antigen in influencing clone behavior by tetramer staining and sorting T cells by the strength of fluorescence signal. They found that high-affinity TCR clones were biased towards a terminal Tex fate while low-affinity TCR clones were biased towards a KLR Tex fate. They ultimately concluded that TCR signal strength directs the phenotypic fate of T cells and that polyclonal T cell responses to chronic antigens allow for a balance of persistence, effector, and memory functions.


  • While the authors suggest that a balance of KLR and terminal Tex may be critical for immune homeostasis, the study lacked further profiling of the functional capabilities of these KLR Tex cells (e.g. ability to produce IFNγ, granzyme B, TNFα). Investigating whether these cells could be reinvigorated by immune checkpoint blockade could also be explored in the future. Further work functionally characterizing the new Tex subsets they identified could add to their discovery by establishing their roles and clinical targetability in different disease contexts.

  • The influence of the tissue microenvironment on Tex was studied by making comparisons across Tex cells isolated from murine, lung, spleen, and liver; however, a limitation of these comparisons was that the red and white pulp of the spleen was not distinguished. These splenic regions have different physiological roles and exposure to antigen, and thus could be differentiated in more detail.

  • Furthermore, it would be valuable to probe publicly available data across cancer or chronic infection disease states to provide evidence of the existence of their newly defined early effector and KLR exhausted T cell phenotypes existing in humans.


With respect to clinical translation, their findings on Tex in chronic LCMV have direct relevance for tumor immunology, where T cell exhaustion can impede several types of cancer immunotherapy. Understanding the gene programs and differentiation pathways underlying these Tex states and how to manipulate them could facilitate improvement of T cell-based immunotherapies. As their work suggests that TCR signal strength directs the phenotypic outcomes of manipulating TCR activation. Additionally, their work suggests that future cellular therapies should aim to promote divergent phenotypes involving both the KLR and terminal Tex states.

On a more philosophic level, their novel findings challenge the current model of T cell exhaustion as a linear progression from progenitor, stem-like Tcf1+Tex cells towards a terminally Tex cell fate. Instead, their new model proposes that TCR signal strength drives the divergent trajectories of exhausted T cells to either a KLR expressing or terminal Tex fate.


Reviewed by Michelle Tran as part of the cross-institutional journal club of the Immunology Institute of the Icahn School of Medicine, Mount Sinai, the Kennedy Institute of Rheumatology and the Oxford Centre for Immuno-Oncology  (OXCIO) (University of Oxford, GB) and Karolinska Institute’s Center for Infectious Medicine (CIM) & Center for Molecular Medicine (CMM).

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