The US16 Gene of Human Cytomegalovirus is Required for Efficient Viral Infection of Endothelial and Epithelial Cells

The human cytomegalovirus (HCMV) US12 gene family comprises a set of 10 contiguous genes (US12 to US21), each encoding a predicted seven-transmembrane protein and whose specific functions have yet to be ascertained. While inactivation of individual US12 family members in laboratory strains of HCMV has not been found to affect viral replication in fibroblasts, inactivation of US16 was reported to increase replication in microvascular endothelial cells. Here, we investigate the properties of US16 further by ascertaining the expression pattern of its product. A recombinant HCMV encoding a tagged version of the US16 protein expressed a 33-kDa polypeptide that accumulated with late kinetics in the cytoplasmic virion assembly compartment. To elucidate the function(s) of pUS16, we generated US16-deficient mutants in the TR clinical strain of HCMV. According to previous studies, inactivation of US16 had no effect on viral replication in fibroblasts. In contrast, the US16-deficient viruses exhibited a major growth defect in both microvascular endothelial cells and retinal pigment epithelial cells. The expression of representative IE, E, and L viral proteins was impaired in endothelial cells infected with a US16 mutant virus, suggesting a defect in the replication cycle that occurs prior to IE gene expression. This defect must be due to an inefficient entry and/or postentry event, since pp65 and viral DNA did not move to the nucleus in US16 mutant-infected cells. Taken together, these data indicate that the US16 gene encodes a novel virus tropism factor that regulates, in a cell-specific manner, a pre-immediate-early phase of the HCMV replication cycle.