Complex roles for proliferating cell nuclear antigen in restricting human cytomegalovirus replication

DNA viruses at once elicit and commandeer host pathways, including DNA repair pathways, for virus replication. Despite encoding its own DNA polymerase and processivity factor, human cytomegalovirus (HCMV) recruits the cellular processivity factor, proliferating cell nuclear antigen (PCNA) and specialized host DNA polymerases involved in translesion synthesis (TLS) to replication compartments (RCs) where viral DNA (vDNA) is synthesized. While the recruitment of TLS polymerases is important for viral genome stability, the role of PCNA is poorly understood. PCNA function in DNA repair is regulated by monoubiquitination (mUb) or SUMOylation of PCNA at lysine 164 (K164). We find that mUb-PCNA increases over the course of infection, and modification of K164 is required for PCNA-mediated restriction of virus replication. mUb-PCNA plays important known roles in recruiting TLS polymerases to DNA, which we have shown are important for viral genome integrity and diversity, represented by structural variants and single nucleotide variants (SNVs), respectively. We find that PCNA drives SNVs on vDNA similar to Y-family TLS polymerases, but this did not require modification at K164. Unlike TLS polymerases, depeletion of PCNA did not result in large-scale rearrangements on vDNA. These striking results suggest separable PCNA-dependent and -independent functions of TLS polymerases on vDNA. By extension, these results imply roles for TLS polymerase beyond their canonical function in TLS in host biology. These findings highlight PCNA as a complex restriction factor for HCMV infection, likely with multiple distinct roles, and provide new insights into the PCNA-mediated regulation of DNA synthesis and repair in viral infection.