PLK1 down-regulates parainfluenza virus 5 gene expression

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.