RecA independent induction of Bacteriophage 434
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Bacteriophage 434 is temperate lambdoid phage that infects Escherichia coli. Similar to the most comprehensively studied bacteriophage lambda (λ), the 434 life cycle alternates between lysogenic and lytic developmental pathways. In the lysogenic pathway, the phage genome is incorporated into the host chromosome, where both are replicated jointly. In the lytic pathway, the phage DNA is not incorporated into the host genome, replicates autonomously, assembles into phages particles and lyses the host cell. Maintenance of the lysogenic state requires the function of the phage repressor protein CI. This DNA binding repressor protein shuts off the transcription of genes necessary for lytic growth, and at the same time directs transcription of its own gene from the integrated phage genome. Host DNA damage induces an SOS system response, in which a host RecA protease induces the CI repressor to autocleave. When autocleaved, the repressor is no longer able to repress the transcription of lytic genes and the lysogenic prophage undergoes lytic growth. During lysogeny, in addition to the CI repressor bacteriophage 434 produces a second protein called HEX. The goals of the experiments described in this dissertation were to determine if changes in the intracellular ionic environment influence the lysis-lysogeny decision, and what function or role does HEX perform in the life cycle of the bacteriophage 434. The results showed that the composition of intracellular ions changes with varying cation concentrations in the growth media. Increased monovalent salt concentrations in the media produced a salt concentration-dependent increase in the frequency of bacteriophage spontaneous induction, which is independent of RecA function. These findings led to the conclusion that in these experiments the increase in phage production is not caused by the SOS pathway; rather, salt stress interferes with the bacteriophage 434 repressor-DNA interactions. Our results also demonstrated that deletions in the phage hex gene increased the spontaneous induction of the bacteriophage 434. Return to wild type levels of spontaneous induction can be accomplished by providing HEX in trans. Furthermore, in vitro studies demonstrated that HEX functions by interfering with RecA mediated autocleavage of the phage CI repressor protein. These findings led us to conclude that HEX protein "fine tunes" the lysis-lysogeny switch of bacteriophage 434.