cDNA cloning of mouse NKR-P1 and genetic linkage with LY-49. Identification of a natural killer cell gene complex on mouse chromosome 6.

NK cells lyse tumor cells and virally infected cells, but the molecular basis for this phenomenon has not been defined. A mAb specific for the rat cell surface molecule, NKR-P1, stimulates rat NK cell lytic activity and is reactive with all rat NK cells, suggesting that this molecule may play a significant role in NK cell function. We have previously described another NK cell-specific Ag, Ly-49, that belongs to a family of cross-hybridizing genes on distal mouse chromosome 6. The rat NKR-P1 Ag shares several features with the mouse Ly-49 Ag, including selective cell surface expression on NK cells, homology to the C-type lectins, expression as a type II integral membrane protein, and disulfide-linked homodimeric structure. To further examine the relationship of NKR-P1 to Ly-49, we have cloned the cDNA encoding a mouse homologue of NKR-P1 (mNKR-P1). The mouse and rat NKR-P1-deduced polypeptide sequences are highly conserved, suggesting a similar tertiary structure. By examination of DNA from informative recombinant inbred mice with Southern blot analysis, we have determined that mNKR-P1 is encoded by a distinct gene that is genetically linked to the Ly-49 locus, lying within 0.5 centi-Morgan (cM) of Ly-49. Although the deduced amino acid sequences of mNKR-P1 and Ly-49 reveal that these proteins are structurally similar, they are only 24% identical at the amino acid level and the cDNA sequences do not demonstrate significant nucleotide homology. Our studies suggest that we have identified a region on mouse chromosome 6 that includes distinct NK-specific genes that encode structurally related proteins (type II integral membrane proteins, C-type lectin super-gene family) but which demonstrate considerable heterogeneity. We have termed this genetic region the NK complex.     

LGL-1: a non-polymorphic antigen expressed on a major population of mouse natural killer cells

Rat mAb have been raised to mouse liver-derived large granular lymphocytes (LGL). One of these mAb (4D11) binds specifically to mouse LGL and appears to recognize a non-allelic determinant on NK-active cell populations. The Ag recognized by 4D11 is expressed on LGL of all mouse strains tested, including C57BL/6 (B6), BALB/c, C3H/HeJ, and SJL/J; thus, we have provisionally called this Ag LGL-1. Analysis of various lymphoid and hemopoietic tissues has indicated that only normal tissues known to contain NK activity have 4D11+ cells. With B6 and B6 congenic strains, a positive correlation exists between the number of LGL in a sample and the percentage of 4D11 immunofluorescence-positive cells detected by flow cytometric analysis. Dual color immunofluorescence analyses indicate that some LGL-1+ cells are also stained for Ly-1 and Thy-1. A very small subset exists that is weakly positive for CD3 and LGL-1. However, virtually no cells are seen which co-express LGL-1 and Ly-2. LU activity against YAC-1 targets was increased 7- to 700-fold in LGL-1+ spleen cells obtained by cell sorting from several different strains of mice (B6, BALB/c, C3H/HeJ, SJL/J, and athymic/nude). Sorted, LGL-1- spleen cells contained little or no NK activity. Cells positively selected for LGL-1 also contained between 50 and 60% LGL by morphology. By using facilitated in vitro antibody plus C' treatments, the majority of NK activity can be depleted from both B6 spleen and liver-derived leukocyte populations enriched for NK cells. mAb 4D11 was also shown to precipitate a protein of approximately 87 kDa from the surface of enriched murine NK cells. This mAb should prove valuable for understanding the role of NK cells in the immune response.     

Monoclonal antibody 4H12 recognizes subsets of adherent-lymphokine activated killer cells and splenic natural killer cells from pregnant and neonatal mice

Using a new mAb, 4H12, that recognizes a plasma membrane-associated Ag on granulated metrial gland cells, we identified subsets of murine NK cells in spleen cell-derived adherent lymphokine activated killer cells and in the spleens of neonatal and pregnant mice. In spleen cell adherent-lymphokine-activated killer cultures, 4H12 Ag was detected on a small subset of cells after 7 days culture and expression increased with time to 70% of the cells after 21 days culture. 4H12+ cells were large (up to 70 microns) and granular. The Ag was also detected in the cytoplasmic granules of some, but not all 4H12 surface positive cells. Coexpression studies indicated 4H12+ cells were predominantly positive for the NK1.1 and ASGM1 Ag, negative for the MAC-1 and F4/80 Ag, and +/- for the LGL-1 and CD3 Ag. Subsets of 4H12+/-, LGL-1+/- exhibited morphologic characteristics restricted to specific phenotypic subsets. The 4H12-/LGL-1+ subset was shown to contain the smallest, least granular cells, whereas the 4H12+/LGL-1+/- subsets contained the largest and most granular cells. Although 4H12 expression was negligible in the spleens of normal adult mice, spleen cells of neonatal and pregnant mice contained subsets of NK1.1+ cells that coexpressed 4H12. The 4H12+/NK1.1+ and 4H12-/NK1.1+ subsets displayed differential levels of NK1.1 expression. 4H12+ NK cells were NK1.1 low to high, whereas 4H12- NK cells were NK1.1 high only. The functional significance of subsets of NK cells in IL-2 culture and in the spleens of neonatal and pregnant mice remains to be elucidated.     

Molecular cloning of the NK1.1 antigen, a member of the NKR-P1 family of natural killer cell activation molecules.

In mice, the NK1.1 alloantigen is expressed on all NK cells and it initiates transmembrane signals that activate cytotoxicity. NK1.1 has been mapped to a chromosomal region that encodes two families of receptor-like molecules, the NKR-P1 family and the Ly-49 family. Both of these gene families encode type II integral membrane proteins whose extracellular domains are homologous with known C-type lectins. We have isolated and expressed a member of the NKR-P1 gene family (mNKR-P1.9) and have demonstrated both by immunofluorescence and by immunoprecipitation that this cDNA encodes NK1.1. These findings, which demonstrate the receptor-like structure of NK1.1, will facilitate studies regarding the role of NK1.1 in natural killing and regarding the identification of possible NK1.1 ligands.     

Stimulation of murine natural killer (NK) cells by a monoclonal antibody specific for the NK1.1 antigen. IL-2-activated NK cells possess additional specific stimulation pathways

NK cells lyse certain tumor cell targets but the effector cell surface molecules responsible for this reactivity remain uncertain. The allotypic NK1.1 Ag is the most specific serologic marker on murine cells that display non-MHC-restricted cytolysis of tumor cell targets, but no function has been previously ascribed to this Ag. In this report, we demonstrate that, in the presence of a mAb specific for the NK1.1 Ag (mAb PK136), freshly isolated and IL-2-activated NK cells from C57BL/6 mice can be induced to lyse an otherwise resistant target cell, Daudi. This phenomenon is effector and mAb specific because NK cells derived from BALB/c mice do not express the NK1.1 Ag and cannot be triggered by mAb PK136. We demonstrate that IL-2 activated but not freshly isolated NK cells express the Ly-6 and VEA Ag, originally described as T cell activation Ag. Moreover, mAb specific for Ly-6 and VEA induce target cell lysis by IL-2 activated but not freshly isolated NK cells. These mAb effects are specific, concentration dependent, and display kinetics that are similar to spontaneous cytolysis of NK-sensitive targets. The Fc portion of the activating antibodies and only FcR bearing target cells participate in mAb-induced activation, consistent with the mechanism of redirected lysis. Finally, analysis of Daudi cells transfected with beta 2-microglobulin gene demonstrate that the expression of MHC class I Ag by the target cell does not affect its sensitivity to mAb-induced lysis by NK cells. These data demonstrate that the NK1.1 Ag is functionally active on both freshly isolated and IL-2-activated NK cells and that IL-2-activated NK cells possess additional pathways of specific stimulation.     

Regulation of the cytotoxic T lymphocyte response against Qa-1 alloantigens

Spleen cells from B6.Tlaa (Qa-1a) mice primed against C57BL/6 (Qa-1b) splenocytes in vivo generate Qa-1-specific CTL when rechallenged with Qa-1b Ag in vitro. The addition of unirradiated Qa-1b splenocytes to these cultures inhibits the generation of Qa-1-specific CTL. By using highly purified cell populations, we demonstrate that the only cell population in resting spleen capable of causing this inhibition is NK1.1+. Although resting CD8 cells lack inhibitory activity, purified CD8 cells precultured with Con A and IL-2 inhibit anti-Qa-1 CTL. This inhibition is specific for the Qa-1b Ag expressed on the inhibitor cells, is not due to cold target competition, and is thus similar to that ascribed to veto cells. Although NK cells from resting spleen inhibit the generation of Qa-1-specific CTL, NK cells precultured in the presence of Con A and IL-2 show an approximate 30-fold increase in veto activity. Thus, NK cells represent the most likely cell population for down-regulating anti-self class I-reactive CTL.     

Mouse NKR-P1B, a novel NK1.1 antigen with inhibitory function

The mouse NK1.1 Ag originally defined as NK cell receptor (NKR)-P1C (CD161) mediates NK cell activation. Here, we show that another member of the mouse CD161 family, NKR-P1B, represents a novel NK1.1 Ag. In contrast to NKR-P1C, which functions as an activating receptor, NKR-P1B inhibits NK cell activation. Association of NKR-P1B with Src homology 2-containing protein tyrosine phosphatase-1 provides a molecular mechanism for this inhibition. The existence of these two NK1.1 Ags with opposite functions suggests a potential role for NKR-P1 molecules, such as those of the Ly-49 gene family, in regulating NK cell function.     

Genomic structure and strain-specific expression of the natural killer cell receptor NKR-P1.

NK cells are able to lyse a variety of virally infected and neoplastic cells in an MHC-unrestricted manner. The cell-surface protein NKR-P1 is thought to play a key role in this process. NKR-P1, initially identified in rat IL-2 activated NK cells, is encoded in the mouse by at least three similar, but not identical, genes. We previously reported the isolation and characterization of three different NKR-P1 cDNA, termed cDNA 2, 34, and 40, from IL-2 activated mouse NK cells. This report describes the structure of the gene encoding NKR-P1 cDNA 2, the smallest of these three cDNA. Gene 2 is composed of six exons spanning approximately 14 kb of genomic DNA. The first exon encodes the N-terminal intracellular domain, and exons 4, 5, and 6 contain the sequences coding for the CRD. This organization is similar to that of other genes that encode C-type animal lectins. The expression of the NKR-P1 genes in A-LAK cells from 13 mouse strains was examined by Northern blot analysis. NKR-P1 expression appears to coincide with that of the NK1.1 Ag. This observation further supports the hypothesis that the NK1.1 Ag is encoded by one of the NKR-P1 genes. Nucleotide sequence analysis of the promoter region of the three NKR-P1 genes in BALB/c and C57BL/6 mice suggests that differences in the level of expression probably do not result from alterations in the upstream regions of these genes, but may be caused by the expression of strain-specific transacting factors.     

Regulation of Ly49D/DAP12 signal transduction by Src-family kinases and CD45

Activating, DAP12-coupled members of the Ly-49 family of NK cell receptors help control viral infections in mice. However, the kinases and/or phosphatases mediating tyrosine phosphorylation of Ly-49D-associated DAP12 have not been elucidated. In this study, we show for the first time that Src family tyrosine kinases are physically and functionally associated with Ly-49D/DAP12 signaling in murine NK cells. Specifically, we demonstrate the following: 1) inhibition of Src family kinases suppresses DAP12 phosphorylation and downstream DAP12 signals; 2) both Fyn and Lck are capable of phosphorylating DAP12; and 3) both kinases coimmunoprecipitate with the Ly-49D/DAP12 complex in NK cells. Although we detect enhanced phosphorylation of Fyn upon Ly-49D cross-linking in NK cells, Ly-49D-mediated events in both Fyn-/- and Fyn/Lck-/- mice appear normal, reinforcing the theme of redundancy in the ability of Src family kinases to initiate activation events. In contrast to disruption of specific Src family enzymes, Ly-49D/DAP12-mediated calcium mobilization and cytokine production by CD45 null NK cells are defective. Although others have ascribed the effects of CD45 mutation solely on the suppression of Src family activity, we demonstrate in this study that DAP12 is hyperphosphorylated in CD45 null NK cells, resulting in uncoordinated tyrosine-mediated signaling upon Ly-49D ligation. Therefore, although our data are consistent with a Src kinase activity proximally within DAP12 signaling, DAP12 also appears to be a substrate of CD45, suggesting a more complex role for this phosphatase than has been reported previously.     

Suppressor T cells and self-tolerance. Active suppression required for normal regulation of anti-erythrocyte autoantibody responses in spleen cells from nonautoimmune mice

Spleen cells from young, nonautoimmune strains of mice cultured with syngeneic E do not develop a significant anti-mouse E response in vitro, consistent with a state of self-tolerance to this Ag. In order to study the role of active suppression in regulating mouse RBC-(MRBC) specific cells in nonautoimmune cell populations, the effect of depleting T cell subsets on the generation of anti-MRBC autoantibodies by nonautoimmune spleen cells was determined. Spleen cells from young BALB/c and C57BL/6 mice were found to generate significant numbers of IgM and IgG anti-MRBC autoantibody-forming cells in culture with MRBC after depletion of Ly-2+ cells by anti-Ly-2 and C treatment. The response which develops is Ag dependent, Ag specific, and dependent upon L3T4+ Th. The magnitude and isotype of this response is similar to the anti-MRBC response generated by spleen cells from 12-mo-old, autoimmune NZB mice and young NZB mice also treated to remove Ly-2+ cells. Addition of isolated Ly-2+ T cells, but not L3T4+ or Ly-2- T cells, to spleen cells depleted of Ly-2+ cells restores apparently normal regulation of the anti-MRBC response in vitro. These data demonstrate that control of a specific autoantibody response to MRBC by nonautoimmune spleen cell populations requires active regulation by an Ly-2+ T cell subset.     

A ligand for the murine NK activation receptor Ly-49D: activation of tolerized NK cells from beta 2-microglobulin-deficient mice

Mouse NK cells express inhibitory NK receptors that recognize target cell MHC class I molecules and activation receptors that are less well defined. The Ly-49D activation receptor on C57BL/6 NK cells recognizes Chinese hamster ovary cells and triggers natural killing. In this study, we demonstrate that a Chinese hamster classical MHC class I molecule is the ligand for Ly-49D in a reporter gene assay system as well as in NK cell killing assays. Ly-49D recognizes the Chinese hamster class I molecule better when it is expressed with Chinese hamster beta(2)-microglobulin (beta(2)m) than murine beta(2)m. However, it is still controversial that Ly-49D recognizes H-2D(d), as we were unable to demonstrate the specificity previously reported. Using this one ligand-one receptor recognition system, function of an NK activation receptor was, for the first time, investigated in NK cells that are tolerized in beta(2)m-deficient mice. Surprisingly, Ly-49D-killing activity against ligand-expressing targets was observed with beta(2)m-deficient mouse NK cells, albeit reduced, even though "tolerized" function of Ly-49D was expected. These results indicate that Ly-49D specifically recognizes the Chinese hamster MHC class I molecule associated with Chinese hamster beta(2)m, and indicate that the Ly-49D NK cell activation receptor is not tolerized in beta(2)m deficiency.     

Mapping of alpha-spectrin on distal mouse chromosome 1

DNAs from different strains of inbred mice and feral Mus spretus were found to exhibit restriction fragment length polymorphisms (RFLP) when hybridized with a probe prepared from a c-DNA clone of the mouse alpha-spectrin (Spna-1) gene. Studies of five recombinant inbred strains and (C57BL/6 X M. spretus) F1 X C57BL/6 backcross mice demonstrated that these RFLPs were allelic and that Spna-1 is closely linked to Ly-9 and Ly-17 on the distal region of chromosome 1.     

Ly-49P activates NK-mediated lysis by recognizing H-2Dd

Little is known regarding the ligand specificity of Ly-49 activating receptor subfamily members expressed by NK cells. A new Ly-49 activating receptor related to Ly-49A in its extracellular domain, designated Ly-49P, was recently cloned from 129 strain mice. We independently cloned an apparent allele of Ly-49P expressed by nonobese diabetic and nonobese diabetes-resistant mouse strain NK cells. We found it to be reactive with the A1 Ab thought to recognize a polymorphic epitope expressed only by the Ly-49A inhibitory receptor of the C57BL/6 strain. Rat RNK-16 cells transfected with Ly-49P mediated reverse Ab-dependent cellular cytotoxicity of FcR-positive target cells, indicating that Ly-49P can activate NK-mediated lysis. We determined that RNK-16 lysis of Con A blasts induced by Ly-49P was MHC dependent, resulting in efficient lysis of H-2Dd-bearing targets. We found that the Dd alpha1/alpha2 domain is required for Ly-49P-mediated RNK-16 activation, as determined by exon shuffling and transfection. Thus, Ly-49P is the second activating Ly-49 receptor demonstrated to induce NK cytotoxicity by recognizing a class I MHC molecule.     

Ly49A allelic variation and MHC class I specificity

The Ly49 family of natural killer (NK) cell receptors is encoded by a polygenic genetic locus. Allelic forms have been described and their expression appears to be regulated. The best-characterized Ly49 molecule, the C57BL/6 form of Ly49A, is an NK cell inhibitory receptor that binds H2Dd. To determine whether differences between Ly49a alleles may have functional consequences, allelic variants of Ly49a were cloned from several inbred mouse strains. Stable transfectants expressing each Ly49a allelic variant were generated and tested for reactivity with a panel of monoclonal antibodies (mAbs A1, JR9.318, YE1/32, and YE1/48) that recognize the C57BL/6 form of Ly49A. Binding to H2Dd was also assessed using fluorescently labeled H2Dd tetramers. Furthermore, cytotoxicity assays were performed using anti-Ly49A mAb-separated interleukin-2-activated NK cells. We show that despite binding to fluorescently labeled H2Dd tetramers, the Ly49A+ NK cells from representative mouse strains displayed significantly different degrees of inhibition with H2Dd targets. These results can be interpreted in the light of recent structural data on the Ly49A-H2Dd complex. Thus, the Ly49 family displays functionally significant allelic polymorphism which adds to the repertoire of NK cell receptors.     

CD73 and Ly-6A/E distinguish in vivo primed but uncommitted mouse CD4 T cells from type 1 or type 2 effector cells

Primed CD4 T cells may develop into effector T cells such as Th1 and Th2, or remain uncommitted as Th primed precursor (Thpp) cells that can subsequently differentiate into Th1 and Th2 cells. Although mouse Thpp-like cells have also been identified among spleen and particularly lymph node cells, further characterization of these cells has been difficult without a defining cell surface marker. Using Affymetrix GeneChips followed by FACS analysis, we found that in vitro-derived Thpp cells expressed CD73 but not Ly-6A/E, whereas Th1 and Th2 cells showed the reciprocal pattern. CD73+ Ly6A/E- memory CD4 T cells were identified in normal C57BL/6 mice, and the proportion of these cells was highest in lymph nodes, lower in spleens, and lowest in the lungs. These cells produced IL-2 and MIP-1alpha, but much less IL-4 and IFN-gamma than CD73- Ly6A/E+ cells. Similar results were obtained with additional Ly-6.2 mouse strains, but not Ly-6.1 strains. Restimulation of Thpp-like CD73+ Ly-6A/E- cells in Th1- or Th2-polarizing conditions induced differentiation into populations producing mainly IFN-gamma or mainly IL-4, respectively. In contrast, the effector-like CD73- Ly-6A/E+ population was more committed, and continued to produce both IL-4 and IFN-gamma in both conditions. CD73 and Ly-6A/E expression therefore identify a population of Thpp-like cells in C57BL/6 mice and at least some other Ly-6.2 mice.     

Allelic variation in the ectodomain of the inhibitory Ly-49G2 receptor alters its specificity for allogeneic and xenogeneic ligands

The Ly-49 multigene receptor family regulates mouse NK cell functions. A number of Ly-49 genes exhibit allelic variation, but the functional significance of allelic differences in extracellular domains of Ly-49 receptors regarding ligand specificity is largely unknown. Amino acid differences exist in the extracellular domains of the B6 and BALB/c allele products of the inhibitory Ly-49G receptor. We constructed chimeric Ly-49 receptors consisting of common cytoplasmic and transmembrane regions of the activating Ly-49W receptor fused with the ectodomains of the B6 and BALB/c alleles of Ly-49G. Expression of these chimeras in the RNK-16 rat NK cell line allowed us to study the specificity of inhibitory receptor ectodomains as they stimulated NK lytic activity. We found that the ectodomain of the BALB/c allele of Ly-49G recognizes both H-2D(d) and D(k) class I MHC alleles, whereas the ectodomain of the B6 allele of Ly-49G recognizes D(d), and not D(k). The specificity for D(k) as well as D(d) of the wild-type Ly-49G(BALB/c) allele product was confirmed with RNK-16 transfectants of this inhibitory receptor. Furthermore, the ectodomain of the Ly-49G(BALB/c) allele recognizes a distinct repertoire of xenogeneic ligands that only partially overlaps with that recognized by Ly-49G(B6). Our results indicate that allelic variation in Ly-49 extracellular domains can have functional significance by altering Ly-49 receptor specificity for mouse class I MHC and xenogeneic ligands.     

Splenic NK1.1-negative, TCR alpha beta intermediate CD4+ T cells exist in naive NK1.1 allelic positive and negative mice, with the capacity to rapidly secrete large amounts of IL-4 and IFN-gamma upon primary TCR stimulation

Splenic NK1.1+CD4+ T cells that express intermediate levels of TCR alpha beta molecules (TCRint) and the DX5 Ag (believed to identify an equivalent population in NK1.1 allelic negative mice) possess the ability to rapidly produce high quantities of immunomodulatory cytokines, notably IL-4 and IFN-gamma, upon primary TCR activation in vivo. Indeed, only T cells expressing the NK1.1 Ag appear to be capable of this function. In this study, we demonstrate that splenic NK1.1-negative TCRintCD4+ T cells, identified on the basis of Fc gamma R expression, exist in naive NK1.1 allelic positive (C57BL/6) and negative (C3H/HeN) mice with the capacity to produce large amounts of IL-4 and IFN-gamma after only 8 h of primary CD3 stimulation in vitro. Furthermore, a comparison of the amounts of early cytokines produced by Fc gamma R+CD4+TCRint T cells with NK1. 1+CD4+ or DX5+CD4+TCRint T cells, simultaneously isolated from C57BL/6 or C3H/HeN mice, revealed strain and population differences. Thus, Fc gamma R defines another subpopulation of splenic CD4+TCRint cells that can rapidly produce large concentrations of immunomodulatory cytokines, suggesting that CD4+TCRint T cells themselves may represent a unique family of immunoregulatory CD4+ T cells whose members include Fc gamma R+CD4+ and NK1.1/DX5+CD4+ T cells.     

Aberrant DAP12 signaling in the 129 strain of mice: implications for the analysis of gene-targeted mice

NK cells are implicated in antiviral responses, bone marrow transplantation and tumor immunosurveillance. Their function is controlled, in part, through the Ly49 family of class I binding receptors. Inhibitory Ly49s suppress signaling, while activating Ly49s (i.e., Ly49D) activate NK cells via the DAP12 signaling chain. Activating Ly49 signaling has been studied primarily in C57BL/6 mice, however, 129 substrains are commonly used in gene-targeting experiments. In this study, we show that in contrast to C57BL/6 NK cells, cross-linking of DAP12-coupled receptors in 129/J mice induces phosphorylation of DAP12 but not calcium mobilization or cytokine production. Consistent with poor-activating Ly49 function, 129/J mice reject bone marrow less efficiently than C57BL/6 mice. Sequence analysis of receptors and DAP12 suggests no structural basis for inactivity, and both the 129/J and C57BL/6 receptors demonstrate normal function in a reconstituted receptor system. Most importantly, reconstitution of Ly49D in 129/J NK cells demonstrated that the signaling deficit is within the NK cells themselves. These unexpected findings bring into question any NK analysis of 129/J, 129Sv, or gene-targeted mice derived from these strains before complete backcrossing, and provide a possible explanation for the differences observed in the immune response of 129 mice in a variety of models.     

Cutting edge: expression of functional CD94/NKG2A inhibitory receptors on fetal NK1.1+Ly-49- cells: a possible mechanism of tolerance during NK cell development.

Fetal liver- and thymus-derived NK1.1+ cells do not express known Ly-49 receptors. Despite the absence of Ly-49 inhibitory receptors, fetal and neonatal NK1.1+Ly-49- cells can distinguish between class Ihigh and class Ilow target cells, suggesting the existence of other class I-specific inhibitory receptors. We demonstrate that fetal NK1. 1+Ly-49- cell lysates contain CD94 protein and that a significant proportion of fetal NK cells are bound by Qa1b tetramers. Fetal and adult NK cells efficiently lyse lymphoblasts from Kb-/-Db-/- mice. Qa1b-specific peptides Qdm and HLA-CW4 leader peptide specifically inhibited the lysis of these blasts by adult and fetal NK cells. Qdm peptide also inhibited the lysis of Qa1b-transfected human 721.221 cells by fetal NK cells. Taken together, these results suggest that the CD94/NKG2A receptor complex is the major known inhibitory receptor for class I (Qa1b) molecules on developing fetal NK cells.     

The effect of the Cmv-1 resistance gene, which is linked to the natural killer cell gene complex, is mediated by natural killer cells.

The resistance of mice to lethal infection by murine CMV (MCMV) is under complex host genetic control with contributions from both H-2 and non-H-2 genes. We have previously shown that an autosomal, non-MHC encoded gene, Cmv-1, controls MCMV replication in the spleen. We have investigated the mechanism by which the Cmv-1 resistance gene confers protection against MCMV infection. Using H-2 compatible irradiation bone marrow chimeras, the enhanced resistance to MCMV infection that is associated with the Cmv-1l allele in the C57BL background was shown to be mediated by an irradiation-sensitive bone marrow-derived cell population, or a factor produced by these cells. The lack of correlation between serum IFN titers and the strain distribution pattern of Cmv-1 in CXB recombinant inbred mouse strains suggests that IFN does not mediate resistance conferred by this gene. Similarly, the lack of effect of in vivo depletion of mature CD4+ and CD8+ T cells on virus replication in C57BL/6J mice indicates that T cells are unlikely to be involved. In contrast, in vivo depletion of NK cells by injection of the anti-NK1.1 mAb PK136 abrogated restricted splenic virus replication in C57BL/6J----BALB.B chimeric mice and in the Cmv-1l CXB strains. These data indicate that the effect of the Cmv-1 gene is mediated by NK cells. The significant augmentation in NK cell activity after MCMV infection of the susceptible Cmv-1h strains (BALB/cBy), CXBG/By, CXBH/By, CXBI/By, and CXBK/By) indicates the existence in these mice of NK cells that are functionally and phenotypically distinct from those in Cmv-1l strains. NK cells present in the Cmv-1h strains are unable to restrict efficiently splenic MCMV replication in vivo, possibly due to a lack of specificity for virus-infected target cells. Finally, flow cytometric analysis of NK1-1 expression in CXB and BXD RI mice together with MCMV replication studies in the BXD RI strains indicate that Cmv-1 is closely linked to NK1.1 and other loci that reside on a distal segment of murine chromosome 6 in a region that has recently been defined as the natural killer complex.