T-bet Fate Mapping Reveals Gestational Stage-Specific Transcriptional Adaptation of Decidual NK Cells

Natural killer (NK) cells are critical regulators of immune balance at the maternal-fetal interface. T-bet (Tbx21) is a key transcription factor shaping NK cell effector functions, yet its role in decidual NK (dNK) cell adaptation across gestation remains unclear.

We used a T-bet fate-mapping mouse model (Rosa26^RFP × Tbx21^Cre) to track developmental and functional reprogramming of NK cells in the uterus, decidua, and placenta throughout pregnancy. Analyses included flow cytometry, bulk RNA sequencing of fate-mapped cells, and single-cell transcriptomic profiling of CD45+Lineage- immune populations at mid and late gestation.

We found that NK cells with a history of T-bet expression (RFP+) progressively downregulate T-bet in a tissue and gestation-specific manner, particularly within decidual and placental compartments. Despite this loss, RFP+ cells retained core NK cell markers and altered their lineage identity towards ILC2 or ILC3 fate. Bulk transcriptomic analysis revealed that T-bet downregulation is associated with dampened IFN-γ, and cytotoxic pathways and increased expression of tissue-residency associated transcriptional regulators. Single-cell RNAseq revealed a gestational transition in dNK subset composition, with a decline in cytotoxic tissue-resident NK cells and expansion of regulatory and conventional NK subsets by late gestation.

These findings identify a novel transcriptional program that shapes NK cell plasticity in response to T-bet downregulation across gestation. Rather than undergoing lineage diversion, dNK cells adapt to the decidual environment via transcriptional compensation and subset redistribution during pregnancy. This work sheds light on the temporal coordination of innate immune function relevant to pregnancy success.