DnaA and LexA Proteins Regulate Transcription of the uvrB Gene in Escherichia coli: The Role of DnaA in the Control of the SOS Regulon

Abstract

The uvrB gene belongs to the SOS network, encoding a key component of the nucleotide excision repair. The uvrB promoter region contains three identified promoters with four LexA binding sites, one consensus and six potential DnaA binding sites. A more than threefold increase in transcription of the chromosomal uvrB gene is observed in both the ΔlexA ΔsulA cells and dnaAA345S cells, and a fivefold increase in the ΔlexA ΔsulA dnaAA345S cells relative to the wild-type cells. The full activity of the uvrB promoter region requires both the uvrBp1-2 and uvrBp3 promoters and is repressed by both the DnaA and LexA proteins. LexA binds tightly to LexA-box1 at the uvrBp1-2 promoter irrespective of the presence of DnaA and this binding is important for the control of the uvrBp1-2 promoter. DnaA and LexA, however, compete for binding to and regulation of the uvrBp3 promoter in which the DnaA-box6 overlaps with LexA-box4. The transcription control of uvrBp3 largely depends on DnaA-box6. Transcription of other SOS regulon genes, such as recN and dinJ, is also repressed by both DnaA and LexA. Interestingly, the absence of LexA in the presence of the DnaAA345S mutant leads to production of elongated cells with incomplete replication, aberrant nucleoids and slow growth. We propose that DnaA is a modulator for maintenance of genome integrity during the SOS response by limiting the expression of the SOS regulon.

H‐NS, IHF, and DnaA lead to changes in nucleoid organizations, replication initiation, and cell division

“…The himA 453, dnaA 345, hns 206 himA 453 mutants grown in ABTGcasa did also not show well‐separated chromosome peaks, and DNA content was less than that in the wild‐type cell (Figure ), again showing that initiation of replication in the mutant cells is asynchronous with delays. The replication pattern of dnaA 345 is in accordance with previous work . It should be noted that the hns 206 mutant contained well‐separated two‐ or four‐chromosome peaks compared with two‐, four‐, or eight‐chromosome peaks in the wild‐type cell (Figure ), indicating that initiation of replication is synchronous but delayed in the hns 206 cells (Figure ).…”

“…Interestingly, a large increase in the number of cells with uncompacted nucleoids and a fraction of cells with abnormal nucleoids were found in the dnaA 345 mutant, and nucleoid compaction caused by DnaA was independent of medium (Figure ), showing that DnaA plays a certain role in nucleoid compaction. A similar finding was also reported previously . Although DnaA has been studied extensively as a replication initiator protein , the function of DnaA on nucleoid compaction has not been investigated thoroughly.…”

“…The DnaA protein initiates chromosomal replication in bacteria . The dnaA 345 mutant is a suppressor for the dnaAN locus and is also found to cause under‐replication and larger cell mass with slower growth . The dnaA 345 mutant and wild‐type cells were exponentially grown in LB or ABTGcasa medium at 37°C and then visualized as described above.…”

“…Also H‐NS can influence the stability of RpoS (σ subunit of RNA polymerase) by inhibiting the expression of IraD and IraM, which are anti‐adapter proteins that stabilize RpoS . DnaA is also a transcription factor, regulating the expression of dnaA , mioC and uvrB . Thus, it is possible that H‐NS and DnaA as transcription factors may modulate nucleoid compaction indirectly by controlling the expression of NAPs.…”

“…The hns or himA gene was amplified by PCR using chromosomal DNA from the wild‐type MG1655 cells as a template and the primers listed in Table . The PCR fragment, including its native promoter, was inserted in pACYC177 at Bam HI and Hin dIII, resulting in plasmid pACYC177‐ hns or pACYC177‐ himA , as described previously . All constructions were sequenced to confirm the plasmid constructions to be correct.…”