Project Summary/Abstract Human Cytomegalovirus (CMV) is a beta-herpesvirus that infects more than half of the US population. Although this persistent/latent infection is asymptomatic in healthy individuals, CMV causes multi-organ diseases in the immune-compromised population (e.g., AIDS patients and solid organ/hematopoietic stem cell transplant recipients). CMV causes not only oral lesions (oral ulceration, periodontal disease, sialadenitis, salivary gland tumor) but also severe systemic diseases (pneumonia, encephalitis, hepatitis, colitis, gastritis). CMV inclusions are frequent in the salivary glands of infants and adults during the course of CMV infection, and oropharyngeal shedding is now a well-established source of human-to-human transmission. We hypothesize that saliva from CMV-infected salivary gland provides immunological impact on the host. In previous reports, human natural killer (NK) cells expressing the activating CD94/NKG2C receptor (heterodimer of CD94 and NKG2C) are expressed in higher frequency in CMV-seropositive adults and preferentially respond during CMV viremia. CD94/NKG2C is known to recognize the invariant human leukocyte antigen (HLA)?E glycoprotein, which is the family of human major histocompatibility complex (MHC) class I molecules. Moreover, HLA-E is abundantly expressed in salivary gland, and soluble form of HLA ligand can be released in a metalloproteinase-dependent fashion. By these facts, we hypothesize that CMV infection alters the peptide repertoire on HLA-E ligand in salivary gland, generating high affinity ligands for the activating CD94/NKG2C receptor (or for the inhibitory CD94/NKG2A receptor), shedding soluble form of HLA-E ligands into saliva, resulting in NK cell activation (or inhibition) through mucosal immune system in oral-pharyngeal cavity (e.g., tonsil and Peyer's patch). To test this hypothesis we propose the two specific aims. Aim 1 will determine the nature of the peptides bound to HLA-E in CMV-infected versus uninfected salivary gland cells by mass spectrometric analysis. Aim 2 will identify CMV-induced HLA-E/peptide complexes that preferentially bind to the activating CD94/NKG2C versus inhibitory CD94/NKG2A receptors. In future research plan, we will assess the ability of candidate peptides to bind and activate primary human oral NK cells. Our studies are intended to determine the role of salivary soluble form of MHC class I molecule as they appear in saliva in the context of CMV infection. Further, from a clinical perspective, the studies we propose are designed to determine whether the HLA-E/peptide complexes deserve investigation as a potential new therapeutic strategy for oral CMV infection.