Jul 2026· Journal of Immunology· Vol 215· 0 citations
TL;DR
It is found that TSC in chronic infection harbor a LEF1+ TSC pool sharing the core stemness epigenetic and molecular program observed in autoimmune LEF1+ TSC, discovering LEF1 as the master regulator defining T cell stemness and identify novel targets for therapeutic intervention.
Abstract
In autoimmunity and chronic infection, both settings of persistent (self or foreign) antigen, immune responses are sustained by stem-like CD8 T cells, which self-renew and give rise to differentiated progeny. However, if and how T stemness is epigenetically encoded, which transcription factor(s) regulate the stem-T cells, and whether the stem-T cell state is disease-specific or shared across diseases, is currently not known.
We used clinically relevant models of autoimmune type 1 diabetes (T1D) and chronic infection and conducted serial T cell transplantation studies in vivo, combined with single cell paired RNA- and ATAC-sequencing on antigen-specific T cells. We developed CRISPR/Cas9-mediated gene-editing approaches in primary T cells as well as CUT&RUN studies, identifying a novel hierarchy of transcription factors regulating stem T cell identity and function.
We discovered that a small subset of stem-T cells (TSC) express lymphoid enhancer-binding factor 1 (LEF1), a member of the TCF/LEF TF family. Paired single cell transcriptomic and epigenomic analyses reveal that the LEF1+ TSC harbor a unique epigenetically encoded molecular state enriched in genes and pathways characteristic of embryonic and adult (somatic) stem cells (e.g. neural stem cells). Strikingly, we found that TSC in chronic infection harbor a LEF1+ TSC pool sharing the core stemness epigenetic and molecular program observed in autoimmune LEF1+ TSC. Loss- and gain-of-function studies in both autoimmune T1D and chronic infection confirmed the critical role of LEF1 in maintaining T cell stemness. CUT&RUN analyses provide clues as to how LEF1 instructs the epigenetically encoded program of stem-T cells.
Here we reveal novel insights into the molecular circuitries of CD8 T cell stemness and differentiation. We discover LEF1 as the master regulator defining T cell stemness and identify novel targets for therapeutic intervention.
NIH R01AI173249, JDRF SRA-2023-1410-S-B, MSKCC Basic Research Innovation Award, The Hearst Foundation
Lymphocyte Differentiation and Peripheral Maintenance (LYM)
Type 1 diabetes (T1D) is a T cell—mediated autoimmune disease driven by β cell-specific CD8 T cells. How autoreactive T cells arise and sustain disease remains unclear. Using the non-obese diabetic mouse model of T1D, we previously identified a stem-like CD8 T cell population in the pancreatic lymph node (pLN) which initiates and sustains β cell destruction: stem T cells (TSC) self-renew and continuously give rise to differentiated progeny (TDIFF) that migrate to the pancreas and kill β cells. Spatial positioning of somatic stem cells is critical for their maintenance, and that niche restricted signals (i.e., WNT and NOTCH) regulate the balance between self-renewal and differentiation. However, if and how T cell stemness is associated with distinct intranodal positioning in pLN, and whether interference in migration disrupts differentiation, is unknown.
We conducted (i) paired single-cell RNA- and ATAC-sequencing, (ii) adoptive T cell transfer studies, (iii) high-resolution imaging, (iv) CRISPR/Cas9 mediated gene editing of autoimmune T cells in pLN to identify the molecular and functional characteristics of TSC and TDIFF.
We discovered unique transcription factors and epigenetic programs governing autoimmune T cell stemness and differentiation. TSC and TDIFF express distinct chemokine receptors and integrins, suggesting that T cell stemness and differentiation are driven by intranodal positioning. Strikingly, WNT and NOTCH signaling were enriched in TSC, driving the expression of critical stem genes, thereby connecting T cell stemness to somatic stem cell biology. Pharmacological blockade and CRISPR/Cas9-mediated deletion of integrins and cell-cell interactions prevented autoimmune T cell differentiation and disease.
Our studies identify novel transcriptional regulators and niche-dependent signals that determine autoimmune T cell stemness and differentiation, opening novel therapeutic avenues for the prevention and treatment of T1D and other autoimmune diseases.
NIH F31DK141119, NIH R01AI173249, Juvenile Diabetes Research Foundation JDRF SRA-2023-1410-S-B, MSKCC Basic Research Innovation Award (BRIA), Hearst Foundation
Lymphocyte Differentiation and Peripheral Maintenance (LYM)
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These findings identify a novel microbial SCFA as a potential metabolic-epigenetic regulator, capable of reactivating effector programs in TEX cells while blocking terminal exhaustion, and provide insights into developing new therapeutic approaches to reprogram TEX cells and enhance the efficacy of T cell immunotherapy.
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Invariant natural killer T (iNKT) cells are innate-like lymphocytes that rapidly respond to lipid antigens presented by CD1d or to inflammatory cytokines and influence diverse immune responses. Much of our current understanding of iNKT cell biology derives from murine studies, which established a framework of thymic differentiation into NKT1, NKT2, and NKT17 subsets. Recent single-cell RNA sequencing (scRNAseq) studies have substantially expanded this view by revealing non-linear developmental trajectories, early epigenomic priming, and a multipotent recent thymic emigrant population that continues to differentiate after thymic egress. In peripheral tissues, iNKT cells undergo extensive remodeling driven by local environmental cues and antigen exposure, giving rise to regulatory, and effector states not observed in the thymus. At the same time, emerging human studies reveal principles that differ from those described in mice. Human iNKT cells exhibit a blended type 1/type 17 transcriptional program, limited evidence for NKT2-like populations, and functionally distinct CD4+, double-negative (DN), CD8αα+, and terminal effector-like subsets. Comparative analyses across species further suggest that differences in thymic selection, transcription factor networks, and peripheral maturation contribute to divergent patterns of iNKT cell specialization. Together, these findings support a revised view of iNKT cells as dynamic and context-dependent transcriptional states shaped by developmental history, tissue environment, antigen exposure, and species-specific regulatory programs.
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Recent studies have identified antigen-experienced TCF1
+
PD-1
+
lineage-negative (lin
-
) CD4 T cells in several tumor types. Strictly, stemness requires durable self-renewal and multipotent output demonstrated at the single-cell level; these criteria have not yet been established for an individual tumor-associated CD4 T cell. Operationally, this review uses “stem-like CD4 T cells” for TCF1
+
PD-1
+
lin
-
populations that lack canonical terminal TH1, TH2, TH17, and Treg commitment programs and show population-level persistence and multilineage output. Pending single-cell lineage tracing, these populations are best regarded as candidate progenitor-like states. Within the tumor microenvironment, their differentiation trajectory may influence antitumor immunity. Treg-mediated suppression, TGF-β signaling, metabolic stress, and limited IL-12 can restrain effector differentiation, whereas release of these constraints can permit TH1 output, CD8-supporting activity in tumor-draining lymph nodes, and direct MHC class II–restricted antitumor effects in selected models. Direct tumor evidence remains concentrated in a limited set of studies, while infection, autoimmunity, transplantation, and CD8 research provide contextual or cross-lineage support. This review therefore presents stem-like CD4 biology as an emerging framework and distinguishes established observations from working models.
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This emerging view frames exhaustion as a context-dependent extension of the memory program rather than its collapse, and highlights how these insights can inform new approaches to manipulate T cell fate for therapeutic benefit.
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