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The superimmunity gene sim of bacteriophage P1 causes superinfection exclusion
1Universität Bielefeld, Fakultät für Biologie, Lehrstuhl für Gentechnologie/Mikrobiologie, Federal Republic of Germany.
Abstract:
Previous work has shown that the sim gene of bacteriophage P1, if cloned into a multicopy vector, confers immunity against P1 infection to cells. We show that a 1.85-kb DNA fragment from the sim region of P1 (in the multicopy plasmid pMK4) expresses immunity and encodes three proteins with molecular weights of about 25, 24, and 15 kDa. Deletion of 650 bp from the sim region abolished synthesis of all three proteins and of the sim phenotype. Expression of sim did not prevent adsorption of P1 to cells. Successful transfection with linear P1 DNA suggests that the recombinational circularization of P1 DNA is not inhibited in the presence of sim. Plasmid pMK4 and a P1 prophage can be stably maintained in the cell indicating that replication of the prophage is not disturbed by sim. The prophage can be induced in the presence of sim. This shows that the sim phenotype is not caused by preventing lytic replication or phage maturation. In cells with pMK4 there is no expression of genes from infecting phages and transduction frequency is drastically reduced. We suggest that sim functions as a superinfection exclusion system by preventing transfer of DNA from the adsorbed phages into the cytoplasm.
Insights
The bacteriophage P1 sim gene confers immunity by preventing infecting phage DNA entry into the cytoplasm. This superinfection exclusion system, mediated by three proteins, blocks gene expression and reduces transduction.
Area of Science:
- Bacteriophage genetics
- Molecular biology
- Microbial genetics
Background:
- The sim gene of bacteriophage P1 is known to confer immunity against P1 infection when present on a multicopy vector.
- Understanding the molecular mechanisms of phage-גן superinfection exclusion is crucial for microbial genetics.
Purpose of the Study:
- To investigate the molecular basis of the sim gene's superinfection exclusion phenotype in bacteriophage P1.
- To identify the proteins encoded by the sim region and their role in conferring immunity.
Main Methods:
- Cloning of a 1.85-kb DNA fragment containing the sim region into a multicopy plasmid (pMK4).
- Deletion analysis of the sim region to assess the role of specific DNA sequences.
- Analysis of protein expression using molecular weights.
- Assessing the effect of sim expression on phage adsorption, DNA transfection, prophage maintenance, induction, and transduction frequency.
Main Results:
- A 1.85-kb fragment from the sim region expressed immunity and encoded three proteins (25, 24, and 15 kDa).
- Deletion of 650 bp from the sim region abolished protein synthesis and the immunity phenotype.
- Sim expression did not inhibit phage adsorption, P1 DNA circularization, prophage replication, or induction.
- Sim expression prevented infecting phage gene expression and drastically reduced transduction frequency.
Conclusions:
- The sim gene functions as a superinfection exclusion system in bacteriophage P1.
- The sim phenotype is mediated by at least three proteins encoded within the 1.85-kb DNA fragment.
- Sim likely prevents superinfection by blocking the transfer of infecting phage DNA into the host cell cytoplasm.