Interactions among Four Proteins Encoded by the Human Cytomegalovirus UL112-113 Region Regulate Their Intranuclear Targeting and the Recruitment of UL44 to Prereplication Foci

Abstract

Four phosphoproteins, of 34, 43, 50, and 84 kDa, with common amino termini are synthesized via alternative splicing from the UL112-113 region of the human cytomegalovirus genome. Although genetic studies provided evidence that both the UL112 and UL113 loci in the viral genome are required for efficient viral replication, whether the four proteins play specific roles or cooperate in replication is not understood. Here we present evidence, using in vitro and in vivo coimmunoprecipitation assays, that the four UL112-113 proteins both self-interact and interact with each other. A mapping study of the 84-kDa protein showed that the N-terminal region encompassing amino acids 1 to 125, which is shared in all UL112-113 proteins and highly conserved among betaherpesviruses, is required for both self-interaction and nuclear localization as foci. Further localization studies revealed that, unlike the 43-, 50-, and 84-kDa proteins, which were distributed as nuclear punctate forms, the 34-kDa form was located predominantly in the cytoplasm. However, when all four proteins were coexpressed simultaneously, all of the UL112-113 proteins were efficiently localized to the promyelocytic leukemia oncogenic domains. We also found that the ability of the UL112-113 proteins to relocate UL44 (the viral polymerase processivity factor) to prereplication foci relied on self-interaction and reached maximal levels when the four proteins were coexpressed. Therefore, our data suggest that interactions occurring among UL112-113 proteins via their shared N-terminal regions are important to both their intranuclear targeting and the recruitment of UL44 to subnuclear sites for viral replication.

Mutations in the M112/M113-Coding Region Facilitate Murine Cytomegalovirus Replication in Human Cells

“…It is highly homologous in its structure to the UL112-113 region of human cytomegalovirus (16,47,60). Although the mechanisms of action of the E1 proteins have not been fully elucidated, a number of studies have shown that they play an important role in the formation of nuclear replication compartments (4,38,40), in viral DNA replication (37,61), and in the regulation of gene expression (24,28,49). Hence, we thought that the mutations in M112/M113 might play a role in the ability of MCMV/h to replicate in human cells.…”

Novel virus-associated proteins encoded by UL112–113 of human cytomegalovirus

“…At the early stages of infection, UL112-113 proteins appear in nuclear domain 10 (ND10) (Ahn et al, 1999;Penfold & Mocarski, 1997). UL112-113 proteins act as pre-replication factors that seed the replication complex by sequentially recruiting single-stranded DNA-binding protein (pUL57) and DNA polymerase processivity factor (pUL44) to ND10 (Park et al, 2006;Penfold & Mocarski, 1997). Furthermore, this observation is strengthened by the fact that UL112-113 interacts with UL44 (Iwayama et al, 1994; Park et al, 2006).…”

A Short cis -Acting Motif in the M112-113 Promoter Region Is Essential for IE3 To Activate M112-113 Gene Expression and Is Important for Murine Cytomegalovirus Replication

“…The M112-113 gene consists of three exons, and alternative splicing creates at least 4 proteins (33,36,38, and 87 kDa) (8,10). UL112-113 is 1 of the 11 core proteins that are essential for HCMV DNA replication (11,12), and it interacts with the lytic origin of replication (13) and forms prereplication domains by recruiting a diversity of viral and cellular proteins that are important for DNA replication (14,15).…”