JC virus-iLOV fluorescent strains enable the detection of early and late viral protein expression

Area of Science:

• Virology
• Molecular Biology
• Drug Discovery

Background:

• John Cunningham virus (JCV) is a prevalent human polyomavirus.
• JCV can cause severe neurological diseases, including progressive multifocal leukoencephalopathy (PML), particularly in immunocompromised individuals.
• Current treatments for JCV infections are lacking, highlighting the urgent need for antiviral therapies, especially for multiple sclerosis patients on immunomodulatory drugs.

Purpose of the Study:

• To develop a novel tool for accelerating the screening of anti-JCV antiviral agents.
• To create a replication-competent JCV strain engineered to express a fluorescent protein.
• To evaluate the utility of fluorescent JCV constructs in drug discovery efforts.

Main Methods:

• Engineered two full-length JCV-iLOV constructs by inserting the improved Light, Oxygen or Voltage-sensing domain (iLOV) gene into the JCV Mad1 backbone.
• Inserted the iLOV gene either before the early regulatory T gene (JCV iLOV-T) or after the late Agno gene (JCV iLOV-Agno).
• Assessed replication competence, fluorescence, viral loads, and pseudovirion production of the engineered JCV strains in vitro.

Main Results:

• Both JCV iLOV strains demonstrated in vitro replication competence and emitted detectable fluorescence.
• The JCV iLOV-T construct exhibited significantly higher cellular and supernatant viral loads compared to JCV iLOV-Agno.
• JCV iLOV-T was capable of producing infectious pseudovirions, indicating its potential for broader applications.

Conclusions:

• The developed JCV iLOV constructs are valuable tools for in vitro drug screening against JCV.
• JCV iLOV-T shows particular promise due to its robust replication and high viral yield.
• These fluorescent reporter systems can significantly simplify and expedite the search for effective anti-JCV therapies.