ICP34.5 Protein of Herpes Simplex Virus Facilitates the Initiation of Protein Translation by Bridging Eukaryotic Initiation Factor 2α (eIF2α) and Protein Phosphatase 1

Abstract

The ICP34.5 protein of herpes simplex virus type 1 is a neurovirulence factor that plays critical roles in viral replication and anti-host responses. One of its functions is to recruit protein phosphatase 1 (PP1) that leads to the dephosphorylation of the ␣ subunit of translation initiation factor eIF2 (eIF2␣), which is inactivated by infection-induced phosphorylation. As PP1 is a protein phosphatase with a wide range of substrates, the question remains to be answered how ICP34.5 directs PP1 to specifically dephosphorylate eIF2␣. Here we report that ICP34.5 not only binds PP1 but also associates with eIF2␣ by in vitro and in vivo assays. The binding site of eIF2␣ is identified at amino acids 233-248 of ICP34.5, which falls in the highly homologous region with human gene growth arrest and DNA damage 34. The interaction between ICP34.5 and eIF2␣ is independent of the phosphorylation status of eIF2␣ at serine 51. Deletion mutation of this region results in the failure of dephosphorylation of eIF2␣ by PP1 and, consequently, interrupts viral protein synthesis and replication. Our data illustrated that the binding between viral protein ICP34.5 and the host eIF2␣ is crucial for the specific dephosphorylation of eIF2␣ by PP1. We propose that herpes simplex virus protein ICP34.5 bridges PP1 and eIF2␣ via their binding motifs and thereby facilitates the protein synthesis and viral replication.Viral infection can activate a series of host immune responses. One of the essential responses is the interruption of the viral protein synthesis, which is executed by doublestranded RNA-dependent protein kinase (PKR) (1). PKR is induced and activated upon viral infection and leads to the phosphorylation of the ␣ subunit of translation initiation factor eIF2 (eIF2␣) 2 at serine 51 (2). The phosphorylation of eIF2␣ globally inhibits the synthesis of viral proteins and cellular proteins (3), thus halting the viral replication.Many viruses have evolved strategies to counteract the antiviral response of PKR (4). For example, PKR activity can be inhibited by HIV-encoded Tar (5), hepatitis C virus NS5A (6), influenza virus NS1 (7), and so forth. In addition to affecting PKR activation as mentioned above, HSV-1 adopted mechanisms not only inhibiting PKR by Us11 (8) but also reversing the biochemical reaction catalyzed by PKR with its neurovirulent factor ICP34.5 (9).ICP34.5 is encoded by the ␥ 1 34.5 gene of HSV-1 and HSV-2. The HSV-1(F) ICP34.5 consists of 263 amino acids and can be divided into three domains: an amino-terminal domain, a linker region of ATP (Ala-Thr-Pro) repeats, and a carboxyl-terminal domain (9, 10). The function of the amino-terminal domain is implicated in the control of TBK1-mediated signaling (11) and also related to autophagy (12). The linker region, with a varying number of the Ala-Thr-Pro repeats, may affect the protein localization (13). The carboxyl-terminal domain is a stretch of 84 amino acids containing a consensus binding motif (R/KVXF) for protein phosphatase 1 (PP1) followed by an Ala-Arg-rich motif and i...

An eIF2α-binding motif in protein phosphatase 1 subunit GADD34 and its viral orthologs is required to promote dephosphorylation of eIF2α

“…However, in support of the scaffolding role of the PP1-targeting subunits, we previously showed that yeast eIF2γ contains a PP1-binding motif that enables recruitment of the phosphatase PP1/ GLC7 to its substrate eIF2α via the eIF2 complex (25). A similar idea has been proposed for the HSV γ34.5 protein, which is able to interact with both PP1 and eIF2α (29).…”

“…Consistent with this idea, deletion of residues 233-248 of the HSV γ34.5 protein (Fig. 3, green box), which shows similarity to GADD34, was shown to impair eIF2α binding (29). To determine whether there is an eIF2α-binding motif in the C-terminal region of GADD34, the amino acid sequences of CReP, γ34.5, and DP71L Fig.…”

“…The consensus PP1-binding RVxF motif and the eIF2α-binding motif are indicated; uppercase letters indicate the most preferred residue in the eIF2α-binding motif. The green box highlights residues that have been shown to be important for the interaction between γ34.5 and eIF2α (29).…”

Time-resolved Global and Chromatin Proteomics during Herpes Simplex Virus Type 1 (HSV-1) Infection

“…ICP34.5 contributes to HSV-1 resistance to the antiviral effects of ␣/␤ interferon (72,73). Moreover, it recruits the serine/threonine-protein phosphatase PPP1CA to dephosphorylate the translation initiation factor EIF2A, thereby counteracting the host shutoff of protein synthesis involving double-stranded RNA-dependent protein kinase EIF2AK2 (74). Recently it has been shown that the interaction between ICP34.5 and C1QBP/p32 at late time points of infection leads to the disintegration of nuclear lamina and facilitates nuclear egress of HSV-1 particles (75).…”

The Us3 Protein of Herpes Simplex Virus 1 Inhibits T Cell Signaling by Confining Linker for Activation of T Cells (LAT) Activation via TRAF6 Protein

“…Us3 interrupts IFN-␤ production by phosphorylation of IRF3 (9). Our previous studies also discovered that the virulent factor ICP34.5 binds with both protein phosphatase 1 and eIF-2␣, facilitating viral protein synthesis (10,11).…”