| Human herpesvirus-7 (HHV-7) is a betaherpesvirus that infects over 90% of the population by the age of 3. Although primary infection is associated with febrile illness, long-term infection with HHV-7 is asymptomatic. In co-evolving with the host immune system, herpesviruses have developed many methods of avoiding detection and clearance during a latent or persistent infection. One common strategy is to interfere with the class I MHC antigen presentation pathway in order to evade detection from cytotoxic T lymphocytes. The HHV-7 U21 protein associates with class I MHC molecules and reroutes them to lysosomes, thereby preventing cell surface expression. The mechanism by which U21 redirects class I molecules to lysosomes is unknown and remains an active area of investigation. In this study, we undergo a thorough characterization of U21’s interaction with class I MHC molecules. We find through bioinformatic analysis that U21 itself may be a structural homolog of class I MHC, and if so is the first example of a virally-encoded class I MHC homolog shown to bind directly to the class I MHC.;We also explore the substrate specificity of U21 for class I MHC and class I MHC-like proteins, finding that U21 associates with both classical and non-classical class I MHC molecules, an unusually broad target range. We test the ability of U21 to associate with class I MHC early in the class I folding pathway, finding that U21 associates with properly-folded class I molecules displaying bound peptide. In vitro analysis reveals that U21 exists in dimeric and tetrameric form in slow equilibrium exchange, with the U21 tetramer capable of binding to class I MHC. U21 associates with class I MHC in a 4:2 ratio, a novel finding for a predicted class I MHC homolog.;We further explore the role of U21’s multimerization in the cell, identifying the putative α3 domain of U21 as sufficient for association with the full-length U21 protein. Finally, we study dominant-negative mutants of U21 to begin to elucidate a model for U21’s multimerization and explore the relationship between U21’s structure and mechanism. This work advances our understanding of the versatility of the class I MHC fold, as well as provides a paradigm for multimerization of class I MHC homologs to allow efficient association with a diverse range of class I MHC proteins. |
