T-bet represses cell cycling in CLL cells and predicts a good outcome. (A) scRNA-seq of Tbx21^–/– and Tbx21^+/+ TCL1 cells was performed and single cells were annotated according to their cell cycle phase. (B) Phospho-specific MS analysis of Tbx21^–/– and Tbx21^+/+ TCL1 cells was performed and kinase networks enriched in Tbx21^–/– TCL1 cells are displayed. (C) The T-bet activity score based on repressed genes was calculated from scRNA-seq data of CLL lymph node samples annotated according to their proliferation status. (D) MEC-1 cell lines with inducible overexpression of T-bet or green fluorescent protein (GFP) as control were generated. Using doxycycline, overexpression was induced and expansion of GFP⁺ control cells and T-bet-overexpressing MEC-1 cells was analyzed using the CellTiter Glo proliferation assay (n = 4 biological replicates á 3 technical replicates). P values were obtained by unpaired t test of the means of biological replicates. (E-G) RNA-seq of patient samples with CLL was performed at diagnosis. Patients with CLL were stratified according to TBX21 mRNA abundance using the maximally selected rank statistics-based cutoff. (E) The OS of TBX21^high vs TBX21^low patients with CLL was analyzed. (F) Patients with CLL were stratified according to TBX21 mRNA abundance using the maximally selected rank statistics-based cutoff and their IGHV mutational status (M = mutated vs UM = unmutated). The OS of M-TBX21^high, M-TBX21^low, UM-TBX21^high, and UM-TBX21^low patients with CLL was analyzed. (G) Patients with CLL were stratified according to TBX21 mRNA abundance using the maximally selected rank and statistics-based cutoff and their ZAP70 gene expression level (ZAP70^high vs ZAP70^low). The OS of ZAP70^high-TBX21^high, ZAP70^high-TBX21^low, ZAP70^low-TBX21^high, and ZAP70^low-TBX21^low patients with CLL was analyzed. P values were obtained by log-rank testing. mRNA, messenger RNA. ∗P ≤ .05.