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Published on: March 5, 2019
PLK1 down-regulates parainfluenza virus 5 gene expression
Dengyun Sun1, Priya Luthra, Zhuo Li
1Intercollege Graduate Program in Cell and Developmental Biology, Pennsylvania State University, University Park, Pennsylvania, USA.
Abstract:
The paramyxoviruses are a family of negative-sense RNA viruses that includes many important human and animal pathogens. Paramyxovirus RNA synthesis requires the viral phosphoprotein (P) and the large (L) protein. Phosphorylation of P is thought to regulate viral gene expression, though direct proof remains elusive. Recently, we reported that phosphorylation of a specific residue (Ser157) of the P protein of parainfluenza virus 5 (PIV5), a prototypical paramyxovirus, correlates with decreased viral gene expression and cytokine expression in infected cells. Here, we show that: Polo-like kinase 1 (PLK1), a serine/theronine kinase that plays a critical role in regulating the cell cycle, interacts with PIV5 P through the S157 residue; PLK1 inhibition increases viral gene expression; PLK1 over-expression inhibits viral gene expression; and PLK1 directly phosphorylates P in vitro, indicating that PLK1 down-regulates viral gene expression by phosphorylating P. Furthermore, we have determined the PLK1 phosphorylation site on P and found that mutant recombinant PIV5 whose P proteins cannot either bind to or be phosphorylated by PLK1 have similar phenotypes. Increased viral gene expression in PIV5 with mutations in the PLK1 binding/phosphorylation sites correlates with increased induction of cell death and cytokine expression, suggesting that PIV5 limits its viral gene expression to avoid these host effects. It is possible that targeting PLK1 will enhance host innate immune responses, leading to a novel strategy of clearing paramyxovirus infections quickly.
Insights
Polo-like kinase 1 (PLK1) down-regulates paramyxovirus RNA synthesis by phosphorylating the viral phosphoprotein (P). Targeting PLK1 may offer a new strategy for treating paramyxovirus infections.
Area of Science:
- Virology
- Molecular Biology
- Immunology
Background:
- Paramyxoviruses are significant human and animal pathogens requiring viral phosphoprotein (P) and large (L) protein for RNA synthesis.
- Phosphorylation of P is hypothesized to regulate viral gene expression, but direct evidence is lacking.
- Previous work linked P phosphorylation at Ser157 in parainfluenza virus 5 (PIV5) to reduced viral and cytokine gene expression.
Purpose of the Study:
- To investigate the role of Polo-like kinase 1 (PLK1) in regulating PIV5 P protein phosphorylation and viral gene expression.
- To identify the interaction and phosphorylation site of PLK1 on PIV5 P.
- To assess the impact of PLK1-P interaction and phosphorylation on viral replication and host response.
Main Methods:
- Co-immunoprecipitation to demonstrate PLK1-PIV5 P interaction.
- In vitro kinase assays to confirm PLK1 phosphorylation of P.
- Site-directed mutagenesis to create PIV5 mutants lacking PLK1 binding/phosphorylation sites.
- Viral gene expression analysis.
- Cell death and cytokine expression assays.
Main Results:
- PLK1 interacts with PIV5 P at the Ser157 residue.
- PLK1 inhibition enhances viral gene expression, while PLK1 overexpression inhibits it.
- PLK1 directly phosphorylates P in vitro, down-regulating viral gene expression.
- PIV5 mutants unable to bind or be phosphorylated by PLK1 exhibit increased viral gene expression, cell death, and cytokine induction.
- PIV5 appears to limit its own gene expression to mitigate host responses.
Conclusions:
- PLK1 negatively regulates PIV5 gene expression through phosphorylation of the P protein.
- PIV5 P protein is phosphorylated by PLK1 at Ser157.
- Mutations disrupting PLK1 binding/phosphorylation of P lead to increased viral replication and host immune responses.
- Targeting PLK1 could be a therapeutic strategy to enhance innate immunity and control paramyxovirus infections.
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