An ELF4 hypomorphic variant results in NK cell deficiency.

NK cell deficiencies (NKD) are a type of primary immune deficiency in which the major immunologic abnormality affects NK cell number, maturity, or function. Since NK cells contribute to immune defense against virally infected cells, patients with NKD experience higher susceptibility to chronic, recurrent, and fatal viral infections. An individual with recurrent viral infections and mild hypogammaglobulinemia was identified to have an X-linked damaging variant in the transcription factor gene ELF4. The variant does not decrease expression but disrupts ELF4 protein interactions and DNA binding, reducing transcriptional activation of target genes and selectively impairing ELF4 function. Corroborating previous murine models of ELF4 deficiency (Elf4-/-) and using a knockdown human NK cell line, we determined that ELF4 is necessary for normal NK cell development, terminal maturation, and function. Through characterization of the NK cells of the proband, expression of the proband's variant in Elf4-/- mouse hematopoietic precursor cells, and a human in vitro NK cell maturation model, we established this ELF4 variant as a potentially novel cause of NKD.

Generation of cell-derived matrices that support human NK cell migration and differentiation.

Human NK cells are effectors of the innate immune system that originate from hematopoietic precursors in the bone marrow. While stromal cell lines that support NK cell development from hematopoietic precursors are often used to generate mature NK cells from lymphoid precursors in vitro, the nature of contributing factors of these stromal cells to the generation of functionally mature NK cells has been poorly described. Previous studies have shown that developing NK cells adhere to, and migrate on, developmentally supportive stroma. Here, we describe the generation of cell-derived matrices (CDMs) from a commonly used murine fetal liver stromal cell line. These CDMs are derived directly from the same EL08.1D2 stromal cell line known to support NK cell differentiation and contain ECM structural components fibronectin and collagen. We demonstrate that CDMs support NK cell adhesion and migration with similar properties as intact cells. Further, we show that CDMs support NK cell maturation from lymphoid precursors in vitro, albeit with reduced cell survival compared to intact cell-based differentiation. Together, these results describe a cell-free system that supports NK cell development and that can serve as a useful model for studying the nature of the biochemical interactions between NK cell developmental intermediates and developmentally supportive substrates.

From stem cell to immune effector: how adhesion, migration, and polarity shape T-cell and natural killer cell lymphocyte development in vitro and in vivo.

Lymphocyte development is a complex and coordinated pathway originating from pluripotent stem cells during embryogenesis and continuing even as matured lymphocytes are primed and educated in adult tissue. Hematopoietic stem cells develop in a specialized niche that includes extracellular matrix and supporting stromal and endothelial cells that both maintain stem cell pluripotency and enable the generation of differentiated cells. Cues for lymphocyte development include changes in integrin-dependent cell motility and adhesion which ultimately help to determine cell fate. The capacity of lymphocytes to adhere and migrate is important for modulating these developmental signals both by regulating the cues that the cell receives from the local microenvironment as well as facilitating the localization of precursors to tissue niches throughout the body. Here we consider how changing migratory and adhesive phenotypes contribute to human natural killer (NK)- and T-cell development as they undergo development from precursors to mature, circulating cells and how our understanding of this process is informed by in vitro models of T- and NK cell generation.

Acquisition of cell migration defines NK cell differentiation from hematopoietic stem cell precursors.

Human natural killer (NK) cells are generated from CD34+ precursors and can be differentiated in vitro by coculture with developmentally supportive stromal cells. We have previously described the acquisition of cell migration as a feature of NK cell terminal maturation in this system. Here we perform continuous long-term imaging and tracking of NK cell progenitors undergoing in vitro differentiation. We demonstrate that NK cell precursors can be tracked over long time periods on the order of weeks by utilizing phase-contrast microscopy and show that these cells acquire increasing motility as they mature. Additionally, we observe that NK cells display a more heterogeneous range of migratory behaviors at later stages of development, with the acquisition of complex modes of migration that are associated with terminal maturation. Together these data demonstrate previously unknown migratory behaviors of innate lymphocytes undergoing lineage differentiation revealed by long-term imaging and analysis workflows.