The Roles of p53, p300, and IFI16 in Human Cytomegalovirus Infection

Abstract

Human Cytomegalovirus (HCMV) is a large double stranded DNA virus. This β-herpesvirus has been implicated in multiple diseases including atherosclerosis, restenosis, and several forms of cancer and is a major cause of disease in unborn children and immunocompromised individuals. This widespread herpesvirus extensively alters the host cell, creating an environment ideal for viral replication. During the virus's characteristically long (72-96 h) replication cycle, it widely regulates cellular processes, driving cells into G1/S phase, blocking apoptosis, and down regulating the immune response. In addition to altering the functions of many host proteins to block unwanted cellular processes, it also targets them to promote viral replication. While there is a large body of evidence describing the role of viral proteins during replication, less is known about the role of host factors. We wanted to determine the role of cellular proteins that were identified through their interactions with viral proteins. In particular, we were interested in IFI16, p53, and p300. Specifically, we hoped to determine: (i) if IFI16, p53, and/or p300 are required for efficient HCMV replication and at what point during the infectious cycle they are needed; and (ii) how the virus modulates and/or alters their function. Using shRNA mediated knockdown, we determined that p53 is not required for efficient replication in fibroblasts, but that p300 and IFI16 are. p300 knockdown resulted in a significant delay in virus release that is likely attributed to defects in early and late RNA and protein accumulation and viral DNA replication. IFI16 is required for expression from the Major Immediate Early Promoter (MIEP) through its interaction with the viral protein pUL83. Interestingly, even though IFI16 does not appear to regulate interferon stimulated genes during HCMV infection, interferon treatment of IFI16 knockdown (IFI16kd) cells further impedes viral replication. Stable expression of pUL83in fibroblasts prior to infection provides some protection against pretreatment with interferon. These results are in line with the immunosuppressive activities already ascribed to pUL83. These data suggest that pUL83 targets IFI16 and, potentially, its interacting partner NFκB to promote expression from the MIEP. Interferon treatment may either disrupt these protein interactions or promotes sequestration of IFI16 away from the MIEP in favor of cellular promoters. It is likely IFI16 may contribute to other viral processes that promote replication.