Temperature-sensitive defect of vesicular stomatitis virus in complementation group II.

Area of Science:

• Virology
• Molecular Biology
• Genetics

Background:

• Vesicular stomatitis virus (VSV) is a model system for studying RNA virus replication.
• Temperature-sensitive (ts) mutants are valuable tools for dissecting viral gene function.
• Complementation group II mutants of VSV exhibit defects in RNA synthesis.

Purpose of the Study:

• To characterize the ts II 052 mutant of VSV.
• To investigate the role of complementation group II in viral RNA synthesis and nucleocapsid formation.
• To identify the specific gene product affected in ts II 052.

Main Methods:

• Infection of HeLa cells with ts II 052 at permissive (30°C) and restrictive (39.5°C) temperatures.
• Analysis of viral RNA synthesis (38S RNA).
• Isolation and characterization of viral nucleocapsids.
• Assessment of nucleocapsid stability (RNase sensitivity, thermal stability, pH stability).

Main Results:

• Reduced viral RNA synthesis and absence of 38S RNA and intracytoplasmic nucleocapsids at the restrictive temperature.
• Nucleocapsid-associated RNA from ts II 052 was sensitive to RNase at the restrictive temperature, unlike wild-type virus.
• Nucleocapsids from ts II 052 lacked the stability of wild-type nucleocapsids when subjected to heat or varying pH.

Conclusions:

• The data indicate an altered interaction between viral RNA and nucleocapsid protein(s) in ts II 052.
• These findings strongly suggest that the complementation group II gene product is the N protein.
• The temperature-sensitive defect in ts II 052 is likely due to an altered N protein, impacting viral RNA synthesis and nucleocapsid stability.