The development and survival of NK cells rely on a complex, spatiotemporal gene expression pattern regulated by specific transcription factors in NK cells and tissue-specific microenvironments supported by hematopoietic cells. Here, we show that somatic deletion of the KLF4 gene, using inducible and lineage-specific cre-transgenic mice, leads to a significant reduction of NK cells (NK1.1(+) TCR-β(-)) in the blood and spleen but not in the BM, liver, or LNs. Functional and immunophenotypic analyses revealed increased apoptosis of CD27(+/-) CD11b(+) NK cells in the spleen of KLF4-deficient mice, although remaining NK cells were able to lyse tumor target cells and produce IFN-γ. A normal recovery of adoptively transferred KLF4-deficient NK cells in WT hosts suggested that the survival defect was not intrinsic of NK cells. However, BM chimeras using KLF4-deficient mice as donors indicated that reduced survival of NK cells depended on BM-derived hematopoietic cells in the spleen. The number of CD11c(hi) DCs, which are known to support NK cell survival, was reduced significantly in the spleen of KLF4-deficient mice, likely a result of a lower number of precDC progenitor cells in this tissue. Taken together, our data suggest that the pluripotency-associated gene KLF4 is required for the maintenance of DCs in the spleen and consequently, survival of differentiated NK cells in this tissue.