Identification and Characterization of Large Granular Lymphocyte (Lgl) Leukemias in F344 Rats

Rodent Ly49 exhibit allele-specific MHC I recognition, yet the interaction site, site 2, encompassing the area below the MHC peptide-binding groove, the α3 domain, and associated β2 microglobulin, is highly conserved among rat and mouse MHC I alleles. We previously demonstrated that allele-specific Ly49 recognition can be affected by polymorphisms specifically in the peptide anchor-binding and supertype-defining B pocket of MHC I, possibly through differential conformations assumed by solvent-exposed interaction residues when articulating with this pocket. Through mutagenesis of RT1-A1c and H-2Dd, we map for the first time the interaction site(s) on rat MHC I mediating rat Ly49i2 recognition and the previously unexamined Ly49GBALB/c interaction with H-2Dd. We demonstrate that rat Ly49i2 and mouse Ly49G use both unique and common interactions at three MHC I H chain subsites to mediate functional binding and allele-specific recognition. We find that the F subsite, formed by solvent-exposed residues below the more conserved C-terminal anchor residue-binding F pocket, acts as an anchoring location for both Ly49i2 and Ly49G, whereas these receptors exhibit distinctive reliance on solvent-exposed residues articulating with the polymorphic anchor-binding and supertype-defining pocket(s) at subsite B, as well as on interaction residues at subsite C in the MHC I α3 domain. Our findings, combined with previous Ly49A/H-2Dd and Ly49C/H-2Kb cocrystal data, suggest how allele-specific MHC I conformations and Ly49 polymorphisms may affect Ly49 placement on MHC I ligands and residue usage at site 2, thereby mediating allele-specific recognition at the highly conserved MHC I interface.

Section: Cell Linesmentioning

confidence: 99%

Distinctive Interactions at Multiple Site 2 Subsites by Allele-Specific Rat and Mouse Ly49 Determine Functional Binding and Class I MHC Specificity

Lavender

Chau

Kane

2007

“…RNK-16 cells were obtained from Dr. C.W. Reynolds (National Cancer Institute, Frederick, MD) (16). The RNK-16.KLRH1 line has been described previously (14).…”

Section: Cell Lines and Reagentsmentioning

confidence: 99%

Identification of an MHC Class I Ligand for the Single Member of a Killer Cell Lectin-like Receptor Family, KLRH1

Daws

Dai

Zinöcker

et al. 2012

Natural killer cells are able to recognize and kill target cells according to differences in MHC class I expression. In rodents, the Ly49 receptors are primarily responsible for this MHC differentiation. We previously described the cloning of a novel C-type lectin–like receptor, KLRH1, encoded in the NK complex adjacent to the Ly49 genes and expressed by subsets of NK and NKT cells. MHC influence on selection of KLRH1+ NK cells in congenic strains suggested that KLRH1 may have an MHC ligand, although we were unable to identify any such ligand. In this study, we have used a sensitive reporter system and Fc fusion protein to demonstrate that KLRH1 binds specifically to the classical MHC class I molecule RT1-A2 of the RT1n haplotype. Cytolytic activity of KLRH1-transfected RNK-16 cells was also inhibited by target cells expressing RT1-A2n. Thus, KLRH1 represents a novel family of MHC allele–specific inhibitory receptors expressed by NK cells.

“…RNK-16, a spontaneous F344 rat strain NK cell leukemia cell line (43), was provided by Dr. M. Nakamura at the University of California (San Francisco, CA), and maintained in RPMI 1640 supplemented with 10% FCS, L-glutamine, penicillin, streptomycin, and 5 ϫ 10 Ϫ5 M 2-ME. RNK-16 effector cells expressing mouse Ly-49 receptors were generated by this laboratory as described (24,25,34) and maintained under G418 selection until 48 h before cytotoxicity assays.…”

Section: Cell Linesmentioning

confidence: 99%

The Rat RT1-A1c MHC Molecule Is a Xenogeneic Ligand Recognized by the Mouse Activating Ly-49W and Inhibitory Ly-49G Receptors

Lavender

Brian

Silver

et al. 2004

Mouse Ly-49 receptors are known to recognize xenogeneic ligands from hamster and rat. However, until now, there has been no description of a specific rat xenogeneic ligand for any mouse Ly-49 receptor. In this report, we identify RT1-A1c, a rat classical class I MHC molecule, as a ligand for the Ly-49GBALB/c inhibitory receptor and the closely related activating receptor, Ly-49W. Xenogeneic class I recognition of targets from PVG but not DA strain rats was mapped to the classical region of the RT1c haplotype by using Con A blasts from RT1c/RT1av1 intra-MHC recombinant rats as targets for RNK-16 cells expressing either Ly-49W or Ly-49GBALB/c receptors. Individual expression of class I molecules from PVG and DA rat strains in YB2/0 target cells demonstrate the xenogeneic recognition to be allele specific, because other class I molecules of the RT1c haplotype, RT1-A2c and RT1-U2c, and a classical class I molecule encoded by the RT1av1 haplotype, RT1-Aa, are not recognized by Ly-49W and -GBALB/c. Furthermore, specificity for RT1-Ac can be transferred from Ly-49W to Ly-49P, which is normally unable to recognize RT1-Ac, by substitution of three residues shared by Ly-49W and -GBALB/c but not Ly-49P. These residues are located in the Ly-49 β4–β5 loop, which can determine class I allele specificity in mouse Ly-49 receptor interactions with mouse class I ligands, suggesting that mouse Ly-49 recognition of rat class I molecules follows similar principles of interaction. These findings have implications for xenotransplantation studies and for discerning Ly-49 recognition motifs present in MHC molecules.