Human cytomegalovirus (HCMV) causes lifelong, persistent infections and its survival is under intense, continuous selective pressure from the immune system. A key aspect of HCMV's capacity for survival lies in immune avoidance. In this context, cells undergoing productive infection exhibit remarkable resistance to natural killer (NK) cell-mediated cytolysis in vitro. To date, six genes encoding proteins (UL16, UL18, UL40, UL83, UL141 and UL142) and one encoding a microRNA (miR-UL112) have been identified as capable of suppressing NK cell recognition. Even though HCMV infection efficiently activates expression of ligands for the NK cell activating receptor NKG2D, at least three functions (UL16, UL142 and miR-UL112) act in concert to suppress presentation of these ligands on the cell surface. Although HCMV downregulates expression of endogenous MHC-I, it encodes an MHC-I homologue (UL18) and also upregulates the expression of cellular HLA-E through the action of UL40. The disruption of normal intercellular connections exposes ligands for NK cell activating receptors on the cell surface, notably CD155. HCMV overcomes this vulnerability by encoding a function (UL141) that acts post-translationally to suppress cell surface expression of CD155. The mechanisms by which HCMV systematically evades (or, more properly, modulates) NK cell recognition constitutes an area of growing understanding that is enhancing our appreciation of the basic mechanisms of NK cell function in humans.

The inhibitory leukocyte Ig-like receptor 1 (LIR-1, also known as ILT2, CD85j, or LILRB1) was identified by its high affinity for the human CMV (HCMV) MHC class I homolog gpUL18. The role of this LIR-1-gpUL18 interaction in modulating NK recognition during HCMV infection has previously not been clearly defined. In this study, LIR-1(+) NKL cell-mediated cytotoxicity was shown to be inhibited by transduction of targets with a replication-deficient adenovirus vector encoding UL18 (RAd-UL18). Fibroblasts infected with an HCMV UL18 mutant (Delta UL18) also exhibited enhanced susceptibility to NKL killing relative to cells infected with the parental virus. In additional cytolysis assays, UL18-mediated protection was also evident in the context of adenovirus vector transduction and HCMV infection of autologous fibroblast targets using IFN-alpha-activated NK bulk cultures derived from a donor with a high frequency of LIR-1(+) NK cells. A single LIR-1(high) NK clone derived from this donor was inhibited by UL18, while 3 of 24 clones were activated. CD107 mobilization assays revealed that LIR-1(+) NK cells were consistently inhibited by UL18 in all tested donors, but this effect was often masked in the global response by UL18-mediated activation of a subset of LIR-1(-) NK cells. Although Ab-blocking experiments support UL18 inhibition being induced by a direct interaction with LIR-1, the UL18-mediated activation is LIR-1 independent.

Natural killer (NK) cells are crucial in the control of cytomegalovirus infections in mice and humans. Here we show that the viral UL141 gene product has an immunomodulatory function that is associated with low-passage strains of human cytomegalovirus. UL141 mediated efficient protection of cells against killing by a wide range of human NK cell populations, including interferon-α-stimulated bulk cultures, polyclonal NK cell lines and most NK cell clones tested. Evasion of NK cell killing was mediated by UL141 blocking surface expression of CD155, which was previously identified as a ligand for NK cell-activating receptors CD226 (DNAM-1) and CD96 (TACTILE). The breadth of the UL141-mediated effect indicates that CD155 has a key role in regulating NK cell function.

Human cytomegalovirus (HCMV) gpUL18 is a HLA class I (HLA-I) homologue with high affinity for the inhibitory receptor LIR-1/ILT2. The previously described 67 kDa form of gpUL18 is shown here to be sensitive to endoglycosidase-H (EndoH). A novel form of gpUL18 with a molecular mass of approximately 160 kDa and resistance to EndoH was identified in cells infected with HCMV strain AD169 or the low passage HCMV isolates Merlin and Toledo. The 67 kDa EndoH-sensitive gpUL18 glycoform was detected earlier in a productive infection (from 24 h post-infection) than the slower-migrating EndoH-resistant glycoform (from 72 h post-infection). Deletion of the US2-US11 region from the HCMV genome was associated with a substantial up-regulation of endogenous HLA-I in infected cells, but had no obvious effect on the gpUL18 expression pattern. Vaccinia virus and adenovirus vectors were used to further analyse gpUL18 expression. Depending on the delivery vector system, differences in the electrophoretic motility of the EndoH-resistant >105 kDa form of gpUL18, but not the EndoH-sensitive 67 kDa form, were observed; post-translational modification of the higher molecular mass glycoform appears to be influenced by active virus infection and vector delivery. The EndoH-sensitive 67 kDa gpUL18 had a rapid turnover, while the maturation to the EndoH-resistant >105 kDa form was relatively slow and inefficient. However, synthesis of the EndoH-resistant >105 kDa form was enhanced with elevated levels of beta2-microglobulin. When expressed by using an adenovirus vector, both the EndoH-sensitive 67 kDa and the EndoH-resistant >105 kDa gpUL18 forms could be detected on the cell surface.