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Cloned bacteriophage phi X174 DNA sequence interferes with synthesis of the complementary strand of infecting
Abstract:
The insertion of a particular phi X DNA sequence in the plasmid pACYC177 strongly decreased the capacity of Escherichia coli cells containing such a plasmid to propagate bacteriophage phi X174. The smallest DNA sequence tested that showed the effect was the HindII fragment R4. This fragment does not code for a complete protein. It contains the sequence specifying the C-terminal part of the gene H protein and the N-terminal part of the gene A protein, as well as the noncoding region between these genes. Analysis of cells that contain plasmids with the "reduction sequence" showed that (i) the adsorption of the phages to the host cells is normal, (ii) in a single infection cycle much less phage is formed, (iii) only 10% of the infecting viral single-stranded DNA is converted to double-stranded replicative-form DNA, and (iv) less progeny replicative form DNA is synthesized. The reduction process is phi X174 specific, since the growth of the related G4 and St-1 phages was not affected in these cells. The effect of the recombinant plasmids on infecting phage DNA shows similarity to the process of superinfection exclusion.
Insights
A specific DNA sequence in Escherichia coli plasmids inhibits bacteriophage phi X174 propagation. This phi X174 DNA fragment reduces phage production by interfering with viral DNA replication, similar to superinfection exclusion.
Area of Science:
- Molecular Biology
- Virology
- Genetics
Background:
- Bacteriophage phi X174 is a well-studied model organism in molecular biology.
- Understanding phage-host interactions is crucial for controlling bacterial infections and developing phage therapy.
Purpose of the Study:
- To investigate the mechanism by which a specific DNA sequence in a plasmid affects bacteriophage phi X174 propagation in Escherichia coli.
- To identify the minimal DNA fragment responsible for this inhibitory effect.
Main Methods:
- Insertion of phi X DNA fragments into the pACYC177 plasmid in Escherichia coli.
- Analysis of phage adsorption, progeny formation, and viral DNA replication in infected cells.
- Testing the specificity of the effect on related phages (G4 and St-1).
Main Results:
- A specific phi X DNA sequence (HindII fragment R4) significantly reduced phi X174 propagation.
- Phage adsorption was normal, but less phage was formed per infection cycle.
- Viral single-stranded DNA to double-stranded replicative-form DNA conversion decreased to 10%.
- Synthesis of progeny replicative form DNA was reduced.
- The effect was specific to phi X174, not affecting G4 or St-1 phages.
Conclusions:
- The identified phi X DNA sequence interferes with early stages of viral DNA replication.
- This interference mechanism resembles superinfection exclusion observed in other phage systems.
- The findings provide insights into phage-host interactions and potential strategies for phage control.