Sequencing of recombinant HCMV AB8

NK cells and T lymphocytes specific for viral antigens presented by classical HLA class Ia molecules play a key role in the control of HCMV infection. Given their reduced polymorphism, HLA-E and -G class Ib molecules were considered unrelated with viral antigen presentation. HLA-E binds conserved nonamers from the leader sequences of other HLA-I molecules, replicated by the UL40 HCMV protein, and interacts with CD94/NKG2 NK cell receptors. Evidence that UL40- and other pathogen-derived peptides may promote HLA-E restricted responses of CD8+ T lymphocytes was reported. By contrast, HLA-G binds a broad spectrum of endogenous sequences but its role in viral antigen presentation is unknown. An experimental approach was set up to search for HCMV antigens displayed by HLA-G in infected cells. Among the analyzed peptidome, 22 sequences corresponding to 16 HCMV molecules were identified; 10 exhibited specific anchor motifs for HLA-G, whose interaction was confirmed in vitro. As compared to the response to IE-1 and pp65 immunodominant antigens detected in short-term (6h) assays, none of the HLA-G-binding peptides stimulated cytokine production by CD8+ T cells from HCMV-seropositive blood donors (n=15). Following a 14-day peptide stimulation of PBMC and expansion with IL-2, CD8+ T cells specifically responding to a subset of these viral antigens were detected in some individuals, however no functional evidence supporting the involvement of HLA-G-restricted T cells was obtained. On the other hand, a subset of viral peptides did bind to both HLA-G and -E but were not recognized by CD94/NKG2 NK cell receptors, with the known exception of HLA-I/UL40 nonamers presented by HLA-E. Our results provide the first evidence that HLA-G may display potentially immunogenic viral peptides in HCMV-infected cells, yet do not support their ability to promote a restricted CD8+ T cell response nor to modulate NK cell functions.