Dissecting the Structural Determinants of the Interaction between the Human Cytomegalovirus UL18 Protein and the CD85j Immune Receptor

Abstract

UL18 is a glycoprotein encoded by the human cytomegalovirus genome and is thought to play a pivotal role during human cytomegalovirus infection, although its exact function is still a matter of debate. UL18 shares structural similarity with MHC class I and binds the receptor CD85j on immune cells. Besides UL18, CD85j binds MHC class I molecules. The binding properties of CD85j to MHC class I molecules have been thoroughly studied. Conversely, very little information is available on the CD85j/UL18 complex, namely that UL18 binds CD85j through its ␣3 domain with an affinity that is ϳ1000-fold higher than the MHC class I affinity for CD85j. Deeper knowledge of features of the UL18/CD85j complex would help to disclose the function of UL18 when it binds to CD85j. In this study we first demonstrated that the UL18␣3 domain is not sufficient per se for binding and that ␤ 2 -microglobulin is necessary for UL18 -CD85j interaction. We then dissected structural determinants of binding UL18 to CD85j. To this end, we constructed a three-dimensional model of the complex. The model was used to design mutants in selected regions of the putative interaction interface, the effects of which were measured on binding. Six regions in both the ␣2 and ␣3 domains and specific amino acids within them were identified that are potentially involved in the UL18 -CD85j interaction. The higher affinity of UL18 to CD85j, compared with MHC class I, seems to be due not to additional interaction regions but to an overall better fit of the two molecules. The Journal of Immunology, 2008, 180: 957-968. T he UL18 is a protein encoded by the human cytomegalovirus (HCMV)4 genome (1). It is a late HCMV Ag, with its transcription occurring from 54 to at least 120 h postinfection (2), and it is not essential for HCMV replication in vitro (3). It shares structural similarity with MHC class I molecules (1). In particular, the extracellular region is formed by three domains (␣1, ␣2, and ␣3) with a sequence identity of 21, 20, and 22% with their class I counterparts (1), respectively. Moreover, its extracellular region associates with ␤ 2 -microglobulin (␤ 2 m) (4) and binds peptides derived from cytoplasmic proteins (5). Similarly to class I molecules, the UL18 ␣2 and ␣3 domains each contains two cysteines characteristic of the MHC Ag-recognition domain fold and of the Ig-like fold, respectively. In contrast, the Ig-like loop of the ␣2 domain is 10 amino acids longer than the corresponding MHC class I loop, and the CD loop of the Ig-like ␣3 domain contains three additional cysteines. As opposed to MHC class I molecules, which have only one to three N-glycosylation sites, the UL18 molecule bears 13 potential glycosylation sites (1), resulting in the expression of highly glycosylated forms (6). The only molecule known so far to bind UL18 is the transmembrane protein CD85j (previously reported as ILT2 or LIR-1) (7).CD85j is a transmembrane receptor expressed on the surface of subsets of NK and T cells and of all B cells and monocytes (7,8). It is formed by four I...

Structure of UL18, a peptide-binding viral MHC mimic, bound to a host inhibitory receptor

“…2), superposition of the UL18 and HLA-A2 ␣1-␣2 domains (Fig. 1C) (rmsd ϭ 0.5 Å for 50 C␣ atoms) revealed that the changes were usually accommodated without generating the protruding loops that had been suggested by computational modeling of the UL18 structure (25,26,28).…”

“…1 A). Both ␣3 domain disulfide bonds are critical for UL18 stability because disruption of either of them abolished the association with ␤2m and the binding to LIR-1 (25,28).…”

Congenital Cytomegalovirus Infection: Molecular Mechanisms Mediating Viral Pathogenesis

“…CMV has evolved several genes that interact with NK-mediated pathways, exploiting both inhibitory and activating pathways. One of the first such identified proteins in HCMV, gpUL18, is encoded by the gene UL18 and appears to exert its NK evasion effect by binding the NK cell inhibitory receptor, LIR-1, with a higher affinity than the host major histocompatability complex (MHC) class I molecule [105-107]. Recently, it has been shown that cells expressing another HCMV MHC class I homolog, the product of the UL142 gene, are protected from NK cell lysis [108].…”

“…The US6 gene encodes a protein that downregulates MHC class I expression [124, 125]. Down-regulation of MHC class I expression could make infected cells susceptible to NK cell mediated-lysis, which (as noted above) is a likely explanation for why CMV encode class I homologs as putative NK evasins [105-107]. …”

Can we build it better? Using BAC genetics to engineer more effective cytomegalovirus vaccines

“…HCMV has evolved several genes that interact with NK cell-mediated pathways, modifying pathways of both inhibition and activation. One such protein, gpUL18, is encoded by the UL18 gene and appears to exert its evasive effect by binding to the NK cell inhibitory receptor leukocyte Ig-like receptor-1 (LIR-1) with a higher affinity than does the host MHC class I molecule (8,9). Recently it has been shown that cells expressing another HCMV MHC class I homolog, the product of the UL142 gene, are protected from NK cell lysis (10).…”