Title: The Control of DNA Replication During Sporulation in Bacillus subtilis
Description: Because of the central role played by genetic information in an organism's ability to survive and reproduce, the genome is stored, copied, and partitioned into daughter cells in a careful, regulated manner. Importantly, the frequency of replication must be regulated so that there are an appropriate number of copies of the genome present in the cell. All organisms accomplish this task by regulating the activity of an initiator protein, which is required to bind to the DNA at the origin of replication for the assembly the replication machinery and the commencement of DNA replication. In bacteria, the initiator protein is DnaA. DnaA experiences positive and negative regulation by both proteins and DNA elements that result in the timely initiation of DNA replication during growth. In response to environmental changes, some bacteria undergo programs of development that result in dramatic changes in morphology and gene expression. One such bacterium in Bacillus subtilis, which produces a dormant and resilient spore in response to nutrient starvation. With a few exceptions, little is known about how DnaA and replication initiation are regulated during bacterial development.

Here, I investigate the regulation of DNA replication during development in B. subtilis. I present evidence that replication is actively inhibited in response to the master regulator of sporulation, SpoOA. I further show that this regulation requires a gene transcribed in the presence of Spo0A, yneE, which I rename sirA, for s[barbelow]porulation i[barbelow]nhibitor of r[barbelow]eplication. The expression of sirA during growth, when it is not usually expressed, results in a growth defect and in the production of cells that lack genetic material. To investigate the mechanism by which sirA inhibits DNA replication, I performed a targeted screen to search for suppressors of sirA expression in the dnaA gene. Four mutations in three amino acids of DnaA allow cells to grow in the presence of SirA. I demonstrate that these residues, which form a patch on the surface of the N-terminal domain of DnaA, make up the interaction site between the DnaA and SirA proteins. Finally, I show that SirA interferes with DnaA's ability to bind the origin of replication.
Subjects: DNA replication