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Defective interfering particles of poliovirus. IV. Mechanisms of enrichment
Abstract:
Infection of HeLa cells by mixtures of standard poliovirus and defective, interfering (DI) poliovirus particles leads to a higher ratio of DI particles in the progeny than in the inoculum. The extent of this enrichment could be varied by various manipulations of the co-infected cells. At any time during the infection cycle, virions made within short times after addition of radioactive uridine were hyperenriched in DI particles; this transient hyperenrichment fell to the equilibrium enrichment level within 45 min after uridine addition. A shift of the temperature of infection from 37 to 31 C also led to a hyperenrichment of DI particles and pulse-labeling revealed a superimposed transient hyperenrichment. By contrast, cells continuously infected at 31 C showed a severe decrement in DI particles apparently because poliovirus DI particles behave as cold-sensitive mutants for RNA synthesis. Cycloheximide treatment early in the infection cycle also led to hyperenrichment. Study of the cycloheximide effect showed that the drug acted as if to change the input ratio of standard to DI particles. These effects on enrichment can be explained as aspects of two different phenomena: enrichment due to preferential DI RNA synthesis and enrichment due to preferential encapsidation of DI RNA. Both mechanisms probably play a role in the normal level of enrichment.
Insights
Infection with defective interfering (DI) poliovirus particles enriches progeny, especially under specific conditions like temperature shifts or cycloheximide treatment. This enrichment results from preferential DI RNA synthesis and encapsidation.
Area of Science:
- Virology
- Molecular Biology
Background:
- Defective interfering (DI) poliovirus particles are variants that require standard poliovirus for replication.
- Co-infection with standard and DI poliovirus leads to an increased proportion of DI particles in the progeny virions compared to the initial mixture.
Purpose of the Study:
- To investigate the mechanisms and conditions that influence the enrichment of DI poliovirus particles during co-infection.
- To understand the roles of DI RNA synthesis and encapsidation in the observed enrichment phenomenon.
Main Methods:
- Infection of HeLa cells with mixtures of standard and DI poliovirus.
- Manipulation of infection conditions, including temperature shifts (37°C to 31°C) and addition of cycloheximide.
- Pulse-labeling experiments with radioactive uridine to track newly synthesized viral RNA and virions.
- Analysis of the ratio of DI particles to standard poliovirus in progeny virions.
Main Results:
- Co-infection resulted in a higher ratio of DI particles in progeny than in the inoculum.
- Transient hyperenrichment of DI particles was observed shortly after radioactive uridine addition, stabilizing within 45 minutes.
- Shifting infection temperature to 31°C caused hyperenrichment, with pulse-labeling revealing a superimposed transient effect.
- Continuous infection at 31°C led to a decrease in DI particles, suggesting cold-sensitive RNA synthesis.
- Early cycloheximide treatment also induced hyperenrichment, mimicking a change in the input ratio of standard to DI particles.
Conclusions:
- The enrichment of DI poliovirus particles is influenced by various experimental conditions.
- Two primary mechanisms contribute to DI particle enrichment: preferential synthesis of DI RNA and preferential encapsidation of DI RNA.
- Both preferential synthesis and encapsidation likely play a role in the overall enrichment observed during poliovirus co-infection.