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Mitosis is required for production of murine leukemia virus and structural proteins during de novo infection
Abstract:
Cloned 3T3FL cells were synchronized in G1 phase of the cell cycle by deprivation of multiplication stimulatory activity of serum and were then infected with Moloney leukemia virus. Eclipse period of virus could be made to vary from less than 10 to 34 h. All virus release was completely dependent and occurred immediately after the first mitosis following serum reconstitution. Virus yield was not affected by the time of virus inoculation as related to the cell DNA synthetic phase. Colchicine arrested the cells in mitosis and prevented the formation of infectious virus. Viral proteins p10, p30, and gp71 were assayed in cell lysates during the growth curve of virus in synchronized cells. The group-specific determinants of each protein were measured in a competition radioimmunoassay. None of the virus proteins appeared during the eclipse period of the virus. All three proteins appeared simultaneously, coincident with mitosis, and continued to rise during the G1 phase. The absolute quantities of each protein were proportional to the amount of Moloney leukemia virus produced. The relative amounts of some of the viral proteins in the cell did not correspond to their content in purified virions suggesting several possible mechanisms of control.
Insights
Moloney leukemia virus replication in synchronized 3T3FL cells is tied to mitosis. Viral protein synthesis and infectious virus release occur post-mitosis, independent of DNA synthesis timing.
Area of Science:
- Virology
- Cell Biology
- Molecular Biology
Background:
- Understanding viral replication dynamics is crucial for developing antiviral strategies.
- The cell cycle influences viral replication, but specific dependencies require elucidation.
- Moloney leukemia virus (MLV) is a retrovirus with implications in oncogenesis.
Purpose of the Study:
- To investigate the relationship between the cell cycle and Moloney leukemia virus replication.
- To determine the timing of viral protein synthesis and infectious virus release in synchronized cells.
- To explore potential regulatory mechanisms controlling viral protein expression.
Main Methods:
- Synchronization of 3T3FL cells in G1 phase using serum deprivation.
- Infection of synchronized cells with Moloney leukemia virus.
- Assay of viral proteins (p10, p30, gp71) using competition radioimmunoassay.
- Analysis of infectious virus release and correlation with cell cycle events.
Main Results:
- The eclipse period of Moloney leukemia virus varied from 10 to 34 hours.
- Infectious virus release was dependent on and occurred immediately after the first mitosis post-serum reconstitution.
- Viral proteins p10, p30, and gp71 appeared simultaneously at mitosis and increased during G1 phase.
- Colchicine-induced mitotic arrest prevented infectious virus formation.
- Absolute viral protein quantities correlated with virus yield, but relative amounts differed from purified virions.
Conclusions:
- Moloney leukemia virus replication and assembly are tightly linked to the host cell's mitotic phase.
- Viral protein synthesis and infectious virus production are initiated at mitosis.
- Discrepancies in viral protein ratios suggest post-transcriptional or post-translational regulatory mechanisms.