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Relationship of retrovirus polyprotein cleavages to virion maturation studied with temperature-sensitive murine
Abstract:
Murine leukemia virus mutants ts3 (Moloney) and ts24 (Rauscher) both formed late-budding structures on the cell membrane at restrictive temperature. They both accumulated core polyproteins Pr65gag and Pr180gag-pol in cell membranes, but the envelope precursor was rapidly turned over. After shift to permissive temperature in the presence of cycloheximide, the accumulated precursors were sequentially cleaved via discrete intermediates both during the final stages of the budding process and in newly released virions to yield the finished virion core proteins and reverse transcriptase. The precursor form of reverse transcriptase was not enzymatically active and became activated partially or entirely inside released virions.
Insights
Murine leukemia virus mutants accumulate viral proteins at the cell membrane. These precursors are processed into active viral components during budding and after release, with reverse transcriptase activation occurring post-release.
Area of Science:
- Virology
- Molecular Biology
- Cell Biology
Background:
- Murine leukemia viruses (MLVs) are retroviruses that cause disease in mice.
- Understanding MLV replication is crucial for developing antiviral therapies.
- Temperature-sensitive mutants provide insights into viral protein processing and assembly.
Purpose of the Study:
- To investigate the assembly and maturation process of Moloney and Rauscher murine leukemia viruses.
- To characterize the processing of viral polyproteins at restrictive and permissive temperatures.
- To determine the enzymatic activity and activation of reverse transcriptase during MLV replication.
Main Methods:
- Utilizing temperature-sensitive mutants (ts3 and ts24) of Moloney and Rauscher MLVs.
- Analyzing viral protein accumulation and localization using cell membrane studies.
- Observing precursor cleavage and virion maturation following temperature shifts.
- Assessing reverse transcriptase activity in precursor and mature forms.
Main Results:
- Mutant MLVs formed late-budding structures at restrictive temperatures.
- Accumulation of core polyproteins (Pr65gag, Pr180gag-pol) in cell membranes was observed.
- Envelope precursor showed rapid turnover, while core precursors were processed upon temperature shift.
- Inactive precursor reverse transcriptase was activated post-virion release.
Conclusions:
- Viral polyprotein processing and maturation are critical steps in MLV replication.
- Sequential cleavage of precursors occurs during late budding and in released virions.
- Reverse transcriptase activation is a late event, occurring after viral release from the host cell.