Abstract
In mice there are two families of MHC class I-specific receptors, namely the Ly49 and CD94/NKG2 receptors. The latter receptors recognize the nonclassical MHC class I Qa-1(b) and are thought to be responsible for the recognition of missing-self and the maintenance of self-tolerance of fetal and neonatal NK cells that do not express Ly49. Currently, how NK cells acquire individual CD94/NKG2 receptors during their development is not known. In this study, we have established a multistep culture method to induce differentiation of embryonic stem (ES) cells into the NK cell lineage and examined the acquisition of CD94/NKG2 by NK cells as they differentiate from ES cells in vitro. ES-derived NK (ES-NK) cells express NK cell-associated proteins and they kill certain tumor cell lines as well as MHC class I-deficient lymphoblasts. They express CD94/NKG2 heterodimers, but not Ly49 molecules, and their cytotoxicity is inhibited by Qa-1(b) on target cells. Using RT-PCR analysis, we also report that the acquisition of these individual receptor gene expressions during different stages of differentiation from ES cells to NK cells follows a predetermined order, with their order of acquisition being first CD94; subsequently NKG2D, NKG2A, and NKG2E; and finally, NKG2C. Single-cell RT-PCR showed coexpression of CD94 and NKG2 genes in most ES-NK cells, and flow cytometric analysis also detected CD94/NKG2 on most ES-NK cells, suggesting that the acquisition of these receptors by ES-NK cells in vitro is nonstochastic, orderly, and cumulative.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Animals
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Antigens, CD / biosynthesis
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Antigens, CD / genetics
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Antigens, CD / metabolism*
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Antigens, CD / physiology
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Cell Differentiation / genetics
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Cell Differentiation / immunology
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Cell Line
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Cells, Cultured
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Embryo, Mammalian
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Histocompatibility Antigens Class I / metabolism
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Humans
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Immunophenotyping
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K562 Cells
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Killer Cells, Natural / cytology*
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Killer Cells, Natural / immunology
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Killer Cells, Natural / metabolism*
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Lectins, C-Type*
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Lymphocyte Subsets / cytology
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Lymphocyte Subsets / immunology
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Lymphocyte Subsets / metabolism
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Membrane Glycoproteins / biosynthesis
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Membrane Glycoproteins / genetics
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Membrane Glycoproteins / metabolism*
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Membrane Glycoproteins / physiology
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Mice
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Mice, Inbred C57BL
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Mice, Knockout
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NK Cell Lectin-Like Receptor Subfamily C
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NK Cell Lectin-Like Receptor Subfamily D
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NK Cell Lectin-Like Receptor Subfamily K
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Receptors, Immunologic / biosynthesis
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Receptors, Immunologic / genetics
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Receptors, Immunologic / metabolism*
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Receptors, Immunologic / physiology
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Receptors, Natural Killer Cell
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Stem Cells / cytology*
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Stem Cells / immunology
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Stem Cells / metabolism*
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Stochastic Processes
Substances
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Antigens, CD
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Histocompatibility Antigens Class I
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KLRC1 protein, human
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KLRC2 protein, human
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KLRC3 protein, human
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KLRD1 protein, human
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KLRK1 protein, human
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Klrc1 protein, mouse
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Klrc2 protein, mouse
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Klrc3 protein, mouse
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Klrd1 protein, mouse
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Klrk1 protein, mouse
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Lectins, C-Type
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Membrane Glycoproteins
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NK Cell Lectin-Like Receptor Subfamily C
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NK Cell Lectin-Like Receptor Subfamily D
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NK Cell Lectin-Like Receptor Subfamily K
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Receptors, Immunologic
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Receptors, Natural Killer Cell