The herpes simplex virus alkaline nuclease is required to maintain replication fork progression

Area of Science:

• Virology
• Molecular Biology
• DNA Replication

Background:

• Herpes simplex virus 1 (HSV-1) is a widespread pathogen causing lifelong infections with potential for severe disease.
• Existing antiviral therapies can be evaded by herpesviruses, necessitating novel therapeutic targets.
• The viral exonuclease UL12 is crucial for producing infectious HSV-1 progeny.

Purpose of the Study:

• To investigate the role of HSV-1 UL12 in viral DNA replication and identify its molecular function.
• To understand the mechanism behind the DNA packaging defect in UL12-deleted HSV-1.
• To explore UL12 as a potential target for antiviral interventions.

Main Methods:

• Isolation of Proteins on Nascent DNA (iPOND) coupled with Stable Isotope Labeling of Amino acids in Cell culture (SILAC) to compare protein abundance at replication forks.
• Quantitative proteomics to analyze protein composition of wild-type (KOS) and UL12-deleted (AN-1) viral replication forks.
• Single-molecule analysis of replication fork progression rates.

Main Results:

• UL12-deleted HSV-1 (AN-1) exhibits aberrant DNA structures preventing capsid packaging.
• Proteomic analysis revealed significant loss of host DNA replication and repair proteins (e.g., MCM2-7, PCNA) at AN-1 replication forks.
• AN-1 replication forks showed decreased progression rates and evidence of stalling, similar to hydroxyurea-treated wild-type forks.

Conclusions:

• HSV-1 UL12 is essential for maintaining DNA replication fork progression.
• The absence of UL12 leads to replication fork stalling and accumulation of structurally abnormal DNA.
• UL12 represents a promising therapeutic target for controlling HSV-1 infections.