High efficiency CRISPR knock-in demonstrates that TCF1 is insufficient to reverse T cell exhaustion

de Menezes, MN; Chen, AXY; Kulkarni, N; Sampurno, S; Saw, NYL; Yap, KM; P&#233;rez-N&#250;&#241;ez, I; Roth, S; Deguit, CDT; Haugen, B; Ramsbottom, KM; Munoz, I; Beavis, PA; Parish, IA

Author(s): de Menezes, MN; Chen, AXY; Kulkarni, N; Sampurno, S; Saw, NYL; Yap, KM; Pérez-Núñez, I; Roth, S; Deguit, CDT; Haugen, B; Ramsbottom, KM; Munoz, I; Beavis, PA; Parish, IA;
Details: Publication Year 2026-02-17,Volume 17,Issue #1,Page 2857
Journal Title: Nature Communications
Publication Type: Research article
Abstract: CD8(+) T cell exhaustion is a regulatory state triggered by chronic antigen stimulation in both cancer and persistent infection. The less differentiated stem-like sub-populations of exhausted T cells have been heavily studied given their importance to the efficacy of current immunotherapies. While the transcription factor TCF1 is both necessary and sufficient for formation and maintenance of these stem-like populations, it remains unclear whether TCF1 can actively de-differentiate more terminally exhausted subsets back into a stem-like state. To address this question, here we utilize and optimize a high efficiency CRISPR knock-in methodology, compatible with mouse in vivo exhaustion models, to engineer T cells that either constitutively over-express TCF1, or conditionally over-express TCF1 following differentiation of the cells into a CX3CR1(+) intermediate-exhausted state. Strikingly, we find that only constitutive, and not conditional, TCF1 over-expression can increase the size of the stem-like T cell pool. Thus, while TCF1 can slow stem-like T cell differentiation, it is insufficient to revert more differentiated cells back into a stem-like state.
Publisher: Springer Nature
Keywords: Animals; *Hepatocyte Nuclear Factor 1-alpha/genetics/metabolism; *Gene Knock-In Techniques/methods; Mice; *CD8-Positive T-Lymphocytes/immunology/metabolism/cytology; Cell Differentiation; Mice, Inbred C57BL; CRISPR-Cas Systems; *T Cell Transcription Factor 1/genetics/metabolism; Mice, Transgenic; *Clustered Regularly Interspaced Short Palindromic Repeats; T-Cell Exhaustion
Department(s): Laboratory Research
Publisher's Version: https://doi.org/10.1038/s41467-026-69671-y
Open Access at Publisher's Site: https://doi.org/10.1038/s41467-026-69671-y
Terms of Use/Rights Notice: Refer to copyright notice on published article.

Creation Date: 2026-03-05 03:10:34

Last Modified: 2026-04-02 06:01:37