Characterization of the FAM111A-Mediated Simian Virus 40 Host Range Phenotype

Abstract

Host range (HR) mutants of simian virus 40 (SV40) with deletions in the C terminus (C-term) of large T antigen (LT) fail to replicate efficiently or form plaques in restrictive cell types. HR mutant viruses exhibit impairments in early and late viral protein expression, DNA replication and virion assembly, although the underlying mechanism for these defects is unknown. Cellular protein FAM111A is an SV40 host range restriction factor that binds directly to LT C-term and whose depletion restores early and late protein expression levels and plaque formation for SV40 HR viruses. FAM111A plays a causative role in two severe human syndromes and has also been implicated in restriction of adenoviruses as well as orthopoxviruses. Lastly, FAM111A has a proposed role in cellular DNA replication, but its cellular and viral restriction functions remain poorly understood. In this Dissertation, we examined the FAM111A-mediated restriction of the SV40 HR phenotype. We found a novel defect of HR viruses in formation of viral replication centers that could be rescued by FAM111A depletion, suggesting that impaired viral replication center formation is a feature of the FAM111A-mediated HR phenotype and may reflect FAM111A function in DNA replication. We also found that FAM111A likely independently restricted viral gene and protein expression and viral replication center formation and that FAM111A also inhibited gene expression from non-viral exogenous DNA, suggesting that FAM111A may play a role in restricting both viral and non-viral exogenous gene expression. Lastly, we observed a nucleolar FAM111A localization pattern during early stages of the cell cycle, followed by dispersal during late G1 and S phase in uninfected cells. In contrast, FAM111A localized to viral replication centers in infection, likely reflective of its proposed role in DNA replication and viral restriction. Collectively, our results indicate that FAM111A restricts gene and protein expression and viral replication center formation for SV40 HR viruses and may function as a broad restriction factor against viral infection and foreign DNA. These findings provide insight into the SV40 HR phenotype and the role of FAM111A in viral restriction and pave the way for uncovering the mechanism underlying the FAM111A contribution to human disease.