Exploring the stage-specific roles of Tcf-1 in T cell development and malignancy at single-cell resolution

Tcf-1 (encoded by Tcf7 ) not only plays critical roles in promoting T cell development and differentiation but also has been identified as a tumor suppressor involved in preventing T cell malignancy. However, the comprehensive mechanisms of Tcf-1 involved in T cell transformation remain poorly understood. In this study, Tcf7 fl/fl mice were crossed with Vav -cre, Lck -cre, or Cd4 -cre mice to delete Tcf-1 conditionally at the beginning of the HSC, DN2–DN3, or DP stage, respectively. The defective T cell development phenotypes became gradually less severe as the deletion stage became more advanced in distinct mouse models. Interestingly, consistent with Tcf7 −/− mice, Tcf7 fl/fl Vav -cre mice developed aggressive T cell lymphoma within 45 weeks, but no tumors were generated in Tcf7 fl/fl Lck -cre or Tcf7 fl/fl Cd4 -cre mice. Single-cell RNA-seq (ScRNA-seq) indicated that ablation of Tcf-1 at distinct phases can subdivide DN1 cells into three clusters (C1, C2, and C3) and DN2–DN3 cells into three clusters (C4, C5, and C6). Moreover, Tcf-1 deficiency redirects bifurcation among divergent cell fates, and clusters C1 and C4 exhibit high potential for leukemic transformation. Mechanistically, we found that Tcf-1 directly binds and mediates chromatin accessibility for both typical T cell regulators and proto-oncogenes, including Myb , Mycn , Runx1 , and Lyl1 in the DN1 phase and Lef1, Id2, Dtx1, Fyn, Bcl11b , and Zfp36l2 in the DN2–DN3 phase. The aberrant expression of these genes due to Tcf-1 deficiency in very early T cells contributes to subsequent tumorigenesis. Thus, we demonstrated that Tcf-1 plays stage-specific roles in regulating early thymocyte development and transformation, providing new insights and evidence for clinical trials on T-ALL leukemia.