Natural killer (NK) cells are lymphocytes that can be distinguished from T and B cells through their involvement in innate immunity and their lack of rearranged antigen receptors. Although NK cells and their receptors were initially characterized in terms of tumor killing in vitro, we have determined that the NK cell activation receptor, Ly-49H, is critically involved in resistance to murine cytomegalovirus in vivo. Ly-49H requires an immunoreceptor tyrosine-based activation motif (ITAM)-containing transmembrane molecule for expression and signal transduction. Thus, NK cells use receptors functionally resembling ITAM-coupled T and B cell antigen receptors to provide vital innate host defense.

Cyclic peptides with an even number of alternating D,L-␣-amino acid residues are known to self-assemble into organic nanotubes. Such peptides previously have been shown to be stable upon protease treatment, membrane active, and bactericidal and to exert antimicrobial activity against Staphylococcus aureus and other gram-positive bacteria. The present report describes the in vitro and in vivo pharmacology of selected members of this cyclic peptide family. The intravenous (i.v.) efficacy of six compounds with MICs of less than 12 g/ml was tested in peritonitis and neutropenic-mouse thigh infection models. Four of the six peptides were efficacious in vivo, with 50% effective doses in the peritonitis model ranging between 4.0 and 6.7 mg/kg against methicillin-sensitive S. aureus (MSSA). In the thigh infection model, the four peptides reduced the bacterial load 2.1 to 3.0 log units following administration of an 8-mg/kg i.v. dose. Activity against methicillin-resistant S. aureus was similar to MSSA. The murine pharmacokinetic profile of each compound was determined following i.v. bolus injection. Interestingly, those compounds with poor efficacy in vivo displayed a significantly lower maximum concentration of the drug in serum and a higher volume of distribution at steady state than compounds with good therapeutic properties. S. aureus was unable to easily develop spontaneous resistance upon prolonged exposure to the peptides at sublethal concentrations, in agreement with the proposed interaction with multiple components of the bacterial membrane canopy. Although additional structure-activity relationship studies are required to improve the therapeutic window of this class of antimicrobial peptides, our results suggest that these amphipathic cyclic D,L-␣-peptides have potential for systemic administration and treatment of otherwise antibiotic-resistant infections.The incidence of community-acquired and nosocomially acquired infections due to the bacterium Staphylococcus aureus is rising (25). From 1990 to 1992, this microorganism was the most common cause of nosocomial pneumonias and surgical wound infections (14). The overall growing crisis in antibiotic resistance and the rise in the incidence of methicillin-resistant S. aureus (MRSA) strains (32, 33) have emphasized the need for therapeutic alternatives to currently available antibiotics. Vancomycin remains the mainstay of therapy against several resistant gram-positive pathogens. However, vancomycin is slowly bactericidal, and with the recent increase in nosocomial infections caused by vancomycin-resistant enterococci and S. aureus (4, 13, 16), there is a growing need for antimicrobial agents with novel mechanisms of action to attack these resistant pathogens. The Food and Drug Administration recently approved the use of daptomycin, a cyclic lipodepsipeptide antibiotic, for the treatment of complicated skin and skin structure infections caused by several gram-positive bacteria. Its mode of action seems to be related to the disruption of the membrane potential of ...

We report the identification of a novel family of genes, named Clr, encoding C-type lectin-like molecules, which maps in the natural killer (NK) gene complex (NKC) on mouse Chromosome 6. Genomic sequence analysis indicates the presence of at least seven members between Nkrpla and Cd69. By RT-PCR, at least three members of the family are expressed on interleukin-2-activated NK cells. Sequence analysis revealed complete open reading frames of 203-205 amino acids, with a carboxyl-terminal C-type lectin-like carbohydrate recognition domain (CRD). The CRDs of the Clr proteins exhibit a significant degree of homology with the known NKC-encoded NK-cell receptors. However, a key cysteine usually present in the CRD is missing in the Clr proteins, suggesting that their ligands and functions are distinct from other molecules encoded in the NKC.

The murine Nkrp1 gene family encodes three previously identified activation and inhibitory receptors expressed on natural killer (NK) cells. This family includes the gene for NKR-P1C (NK1.1), the most specific serologic marker on C57BL/6-derived NK cells and is localized in a gene cluster in the NK gene complex (NKC). To further analyze the Nkrp1 family, we constructed and analyzed a bacterial artificial chromosome contig. A genomic organization of the Nkrp1 family was obtained and three new Nkrp1 genes were isolated from interleukin-2-activated NK cells. Thus, the Nkrp1 family adds to the repertoire of receptors expressed by NK cells.

The NK gene complex (NKC) controls murine cytomegalovirus (MCMV) immunity through Cmv1-dependent natural killer (NK) cell responses. Ly49H expression correlates with Cmv1 phenotypes in different inbred strains, is required for MCMV resistance in C57BL/6 (B6) mice, and its interaction with the MCMV encoded m157 protein leads to NK cell-mediated destruction of virus-infected cells. However, genetic mapping studies have previously indicated that Cmv1 should reside in the D6Wum9-16 NKC interval, distal to Ly49h. Since these data suggested that multiple NKC-linked loci could regulate viral immunity, a putative MCMV resistance control ( Mrc) locus was pinpointed to within the D6Wum9-16 interval on a NKC-aligned bacterial artificial chromosome (BAC). Sequence analysis of BAC 151 revealed several novel G-protein coupled receptor genes, an HMG-1 remnant and many additional polymorphic microsatellites that were useful in determining the minimal genetic interval for the Mrc locus. Moreover, comparison of B6, BALB/c, A/J and recombinant Mrc alleles restricted the genetic interval to approximately 470 bp and showed that it was also a hotspot for recombination. MCMV challenge of novel NKC recombinant mice demonstrated that Mrc(B6) was not required for MCMV resistance nor could it directly complement the Ly49(BALB) haplotype to rescue MCMV susceptibility. Taken together, these data show that while Mrc apparently guides recombination, Ly49H expression is sufficient for MCMV resistance in B6 mice. A direct role for Mrc(B6) in virus resistance is excluded in the novel mice.