Expression of the activating CD94/NKG2C killer lectin-like receptor (KLR) specific for HLA-E was analyzed in peripheral blood lymphocytes (PBLs) from healthy adult blood donors; the expression of other natural killer (NK) cell receptors (ie, CD94/NKG2A, KIR, CD85j, CD161, NKp46, NKp30, and NKG2D) was also studied. Human cytomegalovirus (HCMV) infection as well as the HLA-E and killer immunoglobulin-like receptor (KIR) genotypes were considered as potentially relevant variables associated with CD94/NKG2C expression. The proportion of NKG2C ؉ lymphocytes varied within a wide range (<0.1% to 22.1%), and a significant correlation (r ‫؍‬ 0.83; P < .001) between NKG2C ؉ NK and T cells was noticed. The HLA-E genotype and the number of activating KIR genes of the donors were not significantly related to the percentage of NKG2C ؉ lymphocytes. By contrast, a positive serology for HCMV, but not for other herpesviruses (ie, Epstein-Barr and herpes simplex), turned out to be strongly associated (P < specifically recognize HLA class I molecules and are expressed by natural killer and T-cell subsets. 1-4 Single cells 2 bear variable combinations of these natural killer cell receptors (NKRs), presumably resulting from stochastic gene activation/silencing events that take place during their maturation. 5 The diversity of NKRs observed in different individuals is in part genetically determined, because distinct KIR haplotypes include variable sets of genes. 1 On the other hand, there is evidence that microbial infections may also influence the NKR repertoire. In this regard, murine cytomegalovirus (MCMV) promotes an expansion of NK cells bearing the Ly49H receptor specific for the m157 viral glycoprotein, 6 which plays a crucial role in the immune response to infection. [7][8][9] Moreover, increased proportions of CD8 ϩ T cells with an effector/ memory phenotype bearing inhibitory NKRs (ie, CD94/NKG2A) have been observed in mice infected by different viruses 10-12 as well as in human immunodeficiency virus (HIV)-infected patients. 13 CD94, NKG2A, and NKG2C are C-type lectins encoded at the NK gene complex (NKC) in human chromosome 12. 14 Surface expression of NKG2A/C molecules requires their covalent assembly with CD94. 4,15,16 The CD94/NKG2A heterodimer constitutes an inhibitory receptor that recruits the protein tyrosine phosphatase containing SH2 domain-1 (SHP-1) through the immunoreceptor tyrosine-based inhibitory motif (ITIM)-bearing NKG2A subunit, whereas CD94/NKG2C is coupled to a tyrosine kinase activation pathway through the DAP12 adapter. 17,18 In human beings, both NKRs specifically recognize HLA-E, which presents peptides derived from the signal sequences of other HLA class I molecules [19][20][21][22] ; HLA-E allotypes contain either an Arg (HLA-E R107 ) or a Gly (HLA-E G107 ) at position 107. 23 The biologic relevance of such structural dimorphism remains unclear, but it may affect surface expression levels of the class Ib molecule 24,25 and its interaction with CD94/NKG2 receptors. 26 A number of studies have addre...

Detection of killer-cell immunoglobulin-like receptors (KIR) genes by polymerase chain reaction with sequence-specific primers (PCR-SSP) led in 1997 to the discovery that human genomes diverge largely in the KIR they encode. While only a few KIR genes are conserved in all humans, most individuals lack several those genes, which tend to associate in diverse haplotypic combinations. The PCR-SSP technique, updated to detect the more recently identified KIR genes and alleles, is still used widely to analyze the diversity of human populations, and to study the influence of KIR-gene variability on human health. Several published PCR-SSP methods for KIR genotyping, although simple and robust, have the drawback of relying on the amplification of DNA fragments spanning 0.5-2.0 kbp, which tends to fail in low-quality DNAs. Valuable collections of DNAs often include such poor quality samples, which lead to loss of data and resources. Even worse, undetected falsely negative or positive reactions may result in erroneous gene frequencies and in odd gene combinations. To address those problems, we have redesigned our previously published KIR genotyping method so that it produces short amplicons (less than 200 bp for most genes). This modification minimizes amplification failures, thus conferring greater consistency and reliability to KIR genotyping. In addition, the new PCR-SSP method detects recently described alleles of several KIR genes, and allows for discrimination between the major structural variants of KIR2DS4 and KIR3DP1 without increasing the number of reactions.

Killer-cell Immunoglobulin-like Receptors (KIR) help human natural killer (NK) cells counteract infections by pathogens that evade the immune system by inducing down-regulation of HLA class I molecules in infected cells. KIRs are structural and functionally diverse receptors encoded by a family of polymorphic genes. The most extreme aspect of KIR polymorphism is the varying content of KIR-genes in the genome of different individuals, as first demonstrated by KIR genotyping using the PCR-SSP method. Knowledge on the KIR-gene family has been recently expanded by the identification of new genes, pseudogenes and multiple gene variants, several of which escaped detection by the original genotyping technique. We present here an upgraded PCR-SSP method for KIR genotyping that integrates recent achievements in the research of the diversity of this gene family. Our method permits detection of all known KIR genes and pseudogenes in a 16-reaction set. Furthermore, an additional set of six reactions permits subtyping of KIR2DL5 variants, each of which shows well-differentiated functional and genetic features.

Killer-cell Ig-like receptors (KIR) are structurally and functionally diverse, and enable human NK cells to survey the expression of individual HLA class I molecules, often altered in infections and tumors. Multiple events of non-reciprocal recombination have contributed to the rapid diversification of KIR. We show that *4.5% of the individuals of a Caucasoid population bear a recombinant allele of KIR3DP1, officially designed KIR3DP1*004, that associates tightly with gene duplications of KIR3DP1, KIR2DL4 and KIR3DL1/KIR3DS1. The KIR3DP1 gene is normally silent, but the recombinant allele carries a novel promoter sequence and, as a consequence, is transcribed in all tested individuals. Messenger RNA of KIR3DP1*004 is made up of six exons; of these, exons 1-5 are similar to, and spliced like, those encoding the leader peptide and Ig-domains of KIR3D. By contrast, exon 6 is homologous to no other human KIR sequence, but only to possible homologs in chimpanzees and rhesus macaques, and encodes a short hydrophilic tail. The putative KIR3DP1*004 product, like those of the related genes LAIR-2 and LILRA3/ILT6/LIR4, is predicted to be secreted to the extracellular medium rather than anchored to the cell membrane.