The Bacillus subtilis DinR binding site: redefinition of the consensus sequence

Area of Science:

• Microbiology
• Molecular Biology
• Genetics

Background:

• The SOS response is a critical DNA repair mechanism in bacteria.
• DinR protein was recently identified as the repressor of the SOS response in Bacillus subtilis.
• DinR is thought to bind a consensus sequence in the promoter regions of SOS genes.

Purpose of the Study:

• To investigate the DNA binding characteristics of the DinR protein.
• To define the precise DNA binding site (DinR box) for DinR.
• To understand the mechanism of DinR-mediated repression of the SOS response.

Main Methods:

• Electrophoretic mobility shift assays (EMSA) were used to confirm DinR binding to specific DNA sequences.
• Detailed mutational analysis of the Bacillus subtilis recA operator was performed.
• DNA footprint analysis was employed to determine the boundaries of DinR binding.
• Computational analyses were used to identify potential DinR binding sites in other gram-positive organisms.

Main Results:

• DinR protein binds to consensus sequences upstream of dinA, dinB, dinC, and dinR genes.
• Mutational analysis identified critical nucleotides within the DinR binding site.
• A new consensus sequence for the DinR box (5'-CGAACRNRYGTTYC-3') was established.
• DNA footprinting revealed a 31 bp protected region containing the DinR box.
• DinR binds to the DinR box as a dimer, despite being monomeric in solution.

Conclusions:

• DinR protein functions as a repressor of the SOS response in Bacillus subtilis by binding to the DinR box.
• The DinR box sequence and its central location within a larger protected region are important for DinR binding.
• DinR likely functions as a dimer at the DinR binding site to regulate SOS gene expression.