Defining Roles of UL138 Interactions in the Regulation of pSTAT1 for the Establishment of Latency in Human Cytomegalovirus Infection

Abstract

Innate immune responses are crucial for limiting virus infection. However, viruses often hijack our best defenses for viral objectives. Human Cytomegalovirus (HCMV) is a beta herpesvirus which establishes a life-long latent infection. Defining the virus-host interactions controlling latency and reactivation is vital to the control of viral disease risk posed by virus reactivation. We defined an interaction between UL138, a pro-latency HCMV gene, and the host deubiquintase complexes, UAF1-USP. UAF1 is a scaffold protein pivotal for the activity of ubiquitin specific peptidases (USP), including USP1, USP12, and USP46. UAF1-USP1 sustains an innate immune response through the phosphorylation and activation of signal transducer and activator of transcription-1 (pSTAT1), as well as regulates the DNA damage response. UAF1-USP12 prevents dephosphorylation of pSTAT1 to sustain signaling. After the onset of viral DNA synthesis, pSTAT1 levels are elevated in infection and this depends upon UL138, USP1, and USP12. Upstream kinases, JAK1 and TYK2 are activated regardless of sustained pSTAT1 suggesting UL138-UAF1-USP1 are maintaining pSTAT1 downstream of the kinases. The most likely mechanism of UL138-UAF1-USP1 in maintaining pSTAT1 is deubiquitination of two regulatory monoubiquitin on STAT1 to allow for STAT1 interaction with IFNAR and JAK1 for phosphorylation and activation. Meanwhile, UL138-UAF1-USP12 complexes sustain pSTAT1 by preventing acetylation of pSTAT1 and subsequent dephosphorylation by the phosphatase, PTPN2. Early in infection, pSTAT1 induces specific ISGs but at late times post infection, localizes to viral centers of replication, binds to the viral genome, and influences viral gene expression. Inhibition of USP1 results in a failure to establish latency, marked by increased viral genome replication and production of viral progeny. Inhibition of Jak-STAT signaling also results in increased viral genome synthesis, consistent with a role for UL138-USP1-mediated regulation of STAT1 signaling in the establishment of latency. These findings demonstrate the importance of the UL138-UAF1-USP virus-host interactions in regulating HCMV latency establishment through the control of innate immune signaling. It will be important going forward to differentiate the roles UAF1-USP complexes in latency along with distinguish the purpose of UAF1-USP1 in regulating pSTAT1 relative to its function in the DNA damage response in HCMV infection.