Expression profiles of Bacillus subtilis strains deleted for or ectopically expressing dnaA, sda, and spo0A during exponential growth and after treatment with HPUra.. Bacillus subtilis

The objective of the study was to determine the role of dnaA in regulating gene expression, and the extent to which the effects are mediated by sda and spo0A. Overall design: To identify transcriptional targets for DnaA, we compared a strain lacking dnaA (AIG200) with a strain where dnaA was reintroduced under an IPTG-inducible promoter (Pspank-dnaA; TAW5). Because a dnaA null mutant cannot replicate from oriC, we used an oriC- oriN+ background for these strains. RNA was analyzed from cultures growing exponentially in minimal media, and after treatment with the hydroxyl-phenyl-azo-uracil (HPUra), which inhibits DNA polymerase. In order to determine the effect during exponential growth of deleting sda on gene expression, and the role of sda in mediating the transcriptional effects of dnaA, we analyzed RNA from wild type cells (AG174), an isogenic sda-null (BB668), and the following oriC- oriN+ strains that either lack or inducibly express dnaA and sda: TAW86 (dnaA- Pspank-sda), TAW97 (Pspank-dnaA Pspank-sda), TAW121 (Pspank-dnaA sda-), and TAW118 (dnaA- sda-). Because Sda indirectly inhibits Spo0A, we also analyzed RNA from a spo0A strain (TAW106; Pspank-dnaA, spo0A). For all experiments, at least three biological replicates were performed.