Subject(s)
CD8-Positive T-Lymphocytes/metabolism , Killer Cells, Natural/metabolism , Mast Cells/metabolism , Membrane Proteins/genetics , Proteins , RNA-Binding Proteins/genetics , Animals , Cell Line , Gene Expression , Granulocyte Colony-Stimulating Factor/genetics , Humans , Molecular Sequence Data , Poly(A)-Binding Proteins , Rats , Sequence Analysis , Sequence Analysis, DNA , Sequence Homology, Amino Acid , T-Cell Intracellular Antigen-1ABSTRACT
Three classes of multigene family-encoded receptors enable NK cells to discriminate between polymorphic MHC class I molecules: Ly-49 homodimers, CD94/NKG2 heterodimers and the killer cell inhibitory receptors (KIR). Of these, CD94/NKG2 has been characterized in both rodents and humans. In contrast, Ly-49 family members have hitherto been found only in rodents, and KIR molecules only in the human. In this report, we describe a human cDNA, termed Ly-49L, that constitutes the first human member of the Ly-49 multi-gene family. Compared with rodent Ly-49 molecules, the Ly-49L sequence contains a premature stop codon and predicts a truncated protein that lacks the distal part of a C-terminal lectin domain. Evidence is presented that the premature stop codon results from incomplete excision of the intron between the first two lectin domain exons. Splice variants predicting a full-size Ly-49L protein were not detected. As demonstrated by Northern blot analysis, Ly-49L was transcribed by IL-2-activated NK cells, but not by freshly isolated B or T cells. PCR screening of a 22-clone yeast artificial chromosome contig localized the LY49L locus to the human NK gene complex on chromosome 12p12-p13. Southern blot analysis of genomic DNA showed a simple pattern with a full-length Ly-49L probe at low stringency hybridization conditions, suggesting that Ly-49L may be the only human member of the Ly-49 multigene family.
Subject(s)
Antigens, Ly/genetics , Multigene Family , Alternative Splicing , Amino Acid Sequence , Animals , Antigens, Ly/classification , Base Sequence , Chromosome Mapping , DNA, Complementary , Exons , Humans , Killer Cells, Natural/metabolism , Mice , Molecular Sequence Data , Rats , Sequence Analysis, DNA , Sequence Homology, Amino Acid , Transcription, GeneticABSTRACT
The gene for a rat NK lectin-like receptor (NKLLR), named NKR-P2, has been cloned and characterized. Sequence analysis shows that it represents the orthologue of human NKG2D and that the two molecules form a distinct NKLLR family, no more related to NKG2A/B, -C or -E than to other NKLLR families. Nkrp2 is a single-copy gene containing seven introns, mapping to the rat NK gene complex. Rat NKR-P2 differs from the human orthologue in that its cytoplasmic tail contains 13 additional amino acids, encoded by a separate exon. Splice variants lacking this exon were not detected in T cells or NK cells. NKR-P2 is strongly expressed by NK cells. In contrast to other NKLLR, it is also strongly expressed by resting thoracic duct CD4+ and CD8+ T cells, but not by thymocytes or other hemopoietic cells.
Subject(s)
CD4-Positive T-Lymphocytes/ultrastructure , CD8-Positive T-Lymphocytes/ultrastructure , Killer Cells, Natural/ultrastructure , Lectins, C-Type , Lectins/metabolism , Receptors, Immunologic/biosynthesis , Receptors, Immunologic/genetics , Alternative Splicing , Amino Acid Sequence , Animals , Chromosome Mapping , Cloning, Molecular , DNA, Complementary/genetics , Exons , Introns , Molecular Sequence Data , NK Cell Lectin-Like Receptor Subfamily K , Rats , Rats, Inbred F344 , Sequence Homology, Amino AcidABSTRACT
Two different lectin-like receptors for MHC class I molecules have so far been identified on natural killer (NK) cells, the Ly-49 homodimeric receptors in mice and the NKG2/CD94 heterodimeric receptors in humans. The recent identification of a rat CD94 orthologue implied that NK cell receptors equivalent to NKG2/CD94 also exist in rodents. Here we describe the cDNA cloning of two rat genes homologous to members of the human NKG2 multigene family. The deduced rat NKG2A protein contains a cytoplasmic immunoreceptor tyrosine-based inhibition motif (ITIM), whereas the cytoplasmic tail of rat NKG2C lacks ITIM. The genes map to the rat NK gene complex and are selectively expressed by NK cells. The expression is strain dependent, with high expression in DA and low in PVG NK cells, correlating with the expression of rat CD94. Ly-49 genes have previously been identified in the rat, and the existence of rat NKG2 genes in addition to a CD94 orthologue suggests that NK cell populations utilize different C-type lectin receptors for MHC class I molecules in parallel.
Subject(s)
Antigens, CD/genetics , Killer Cells, Natural/metabolism , Lectins, C-Type , Membrane Glycoproteins/genetics , Sequence Homology, Amino Acid , Sequence Homology, Nucleic Acid , Amino Acid Sequence , Animals , Antigens, CD/biosynthesis , Antigens, Surface/genetics , DNA, Complementary/isolation & purification , Humans , Membrane Glycoproteins/biosynthesis , Molecular Sequence Data , NK Cell Lectin-Like Receptor Subfamily B , NK Cell Lectin-Like Receptor Subfamily C , NK Cell Lectin-Like Receptor Subfamily D , Polymorphism, Restriction Fragment Length , Rats , Receptors, Immunologic/genetics , Receptors, Natural Killer Cell , Sequence Analysis, DNAABSTRACT
Three classes of major histocompatibility (MHC) class I binding receptors on natural killer (NK) cells have so far been described: CD94/NKG2 heterodimeric receptors and killer cell inhibitory receptors in the human, and Ly-49 homodimers in rodents. CD94, NKG2 and Ly-49 belong to the C-type lectin superfamily. As yet, CD94 and NKG2 molecules have not been detected in rodents or Ly-49 in humans. It has therefore been proposed that the two receptors represent functional equivalents in these species. The present study describes the cDNA cloning of a novel rat gene encoding a protein of 179 amino acids, 54.2% identical to human CD94. The single-copy Cd94 gene is localized to the rat NK gene complex (NKC), within 50 kb from Nkrp2, between the Nkrp1 and Ly49 gene clusters. By Northern blot analysis, we showed that rat CD94 is selectively expressed by NK cells and a small subset of T cells, similar to the human orthologue. This expression is strain dependent, with high expression in DA NK cells and low in PVG NK cells. Evidence is presented that this difference is not due to receptor repertoire shaping by MHC-encoded ligands, but is controlled by genetic elements residing within the NKC. The identification of a rat CD94 orthologue suggests that NK cell populations utilize two different C-type lectin receptors for MHC class I molecules in parallel.
