Background Herpes simplex virus types 1 and 2 (HSV-1 and HSV-2) are among the most prevalent human pathogens in both industrialised and developing regions. In the current study, we reported that two zinc ionephores, pyrrolidine dithiocarbamate (PDTC) and pyrithione (PT) could inhibit herpes simplex virus type 1 and type 2 (HSV-1 and HSV-2) replication.
Methods Viral replication was evaluated via detecting HSV late gene product (Glycoprotein, gD) using In-cell Western. NF-κB, MAPK activation, viral immediate-early genes expression and profile of cellular ubiquitin-conjugates were determined by Western blot. Protein localization was investigated via immunofluorescence staining. Viral protein expression on mRNA level was quantified by realtime-PCR. 26S Proteasome activation was determined using fluorogenic substrate, Suc-LLVY-AMC.
Results PDTC and PT inhibited HSV-1 and HSV-2 gD expression and the production of viral progeny, which was dependent on zinc ion. Further studies showed that these two compounds suppressed the HSV immediate-early gene, the infected cell polypeptide 4 (ICP4) expression, but had less effect on ICP0. HSV infection could interact with cellular ubiquitin-proteasome system (UPS) and cause loss of high molecular weight ubiquitin-conjugates. It was found that PDTC and PT could interfere UPS, leading to the inhibition of HSV-2-induced IκB-α degradation to prevent NF-κB activation and enhanced PML stability in nucleus. However, PDTC and PT did not show direct inhibition of 26S proteasome activity. Instead, these two zinc ionphores induced import of extracellular zinc ions into cells, which facilitated dysregulation of proteasome function and accumulation of intracellular ubiquitin-conjugates. Other evidence was that the inhibitors of ubiquitin activating enzyme E1 and deubiquitinase also inhibited HSV replication, implying that UPS was required for effective replication of HSV-1 and HSV-2
Conclusion Homostasis of ubiquitin cycle and UPS were critical for HSV gene expression and replication and that the ubiquitin-proteasome pathway is a potential drug target for HSV infection.
- herpes simplex virus
- ubiquitin-proteasome system
- Zinc ionophore
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