The RNA- and DNA-Binding Protein Y-Box Binding Protein 1 (YB-1) Regulates iNKT Cell Development

Y-box binding protein 1 (YB-1) is a multifunctional RNA- and DNA-binding protein with broad regulatory functions in gene expression, particularly at the posttranscriptional level. Here, we demonstrate that conditional deletion of YB-1 at the double-positive (DP) thymocyte stage causes an ∼80% reduction of invariant natural killer T (iNKT) cells in thymus, spleen, and liver, evident already in day-14 neonates and persisting into adulthood. Our data reveal CD44^-NK1.1^- stage 1 accumulation and a selective loss of CD44⁺NK1.1⁺ stage 3 iNKT cells, indicating a postselection maturation defect. All iNKT cell subsets (iNKT1, iNKT2, iNKT17) were reduced, with thymic iNKT1 and splenic iNKT17 cells most severely affected. PMA/ionomycin-stimulated YB-1-deficient iNKT cells showed preserved IFN-γ⁺/IL-4⁺ frequencies but reduced per-cell cytokine production and a loss of IL-17 production. Interestingly, YB-1^KO DP thymocytes showed increased CD1d levels, suggesting increased TCR signal strength in the thymus of YB-1-deficient mice. Whereas CD5 levels were elevated, basal Nur77, ICOS, and CD122 (IL-15Rβ) expression were reduced in iNKT cells. Furthermore, apoptosis was increased, particularly at iNKT stages 2-3. Together, these findings identify YB-1 as a central regulator of iNKT cell development that integrates TCR, co-stimulatory, and IL-15 signaling to ensure postselection iNKT cell maturation, effector subset specification, and survival.