Interaction of a repressor and its binding sites for regulation of the Bacillus subtilis iol divergon

Transcription of the Bacillus subtilis iol divergon is negatively regulated by a repressor encoded by iolR, which belongs to the DeoR family of bacter ial regulators. Gel retardation analysis involving the IolR protein synthes ized in Escherichia coli revealed that IolR bound specifically and independ ently to each of the iol and iolRS promoter regions, with higher affinity t o iol. DNase I footprinting revealed that IolR affected DNase I sensitivity either in the iol promoter region between nucleotides -46 and +51 or in io lRS between -79 and -2 (+1 is the transcription initiation nucleotide of bo th iol and iolRS), indicating its interaction with the extended regions of the iol and iolRS promoters. Deletion analysis indicated that the iol regio n between -23 and +21 is involved mainly in IolR binding and negative regul ation, while the iolRS region between -70 and -44 comprises at least part o f the cis-acting sequences for IolR binding and negative regulation. Sequen ce examination of the extended regions revealed that a tandem direct repeat consisting of two relatively conserved Il-mer sequences, WRAYCAADARD (wher e D is A, G or T; R is A or G; W is A or T; and Y is C or T), found in each of the iol and, iolRS regions might be a determinant sequence for the IolR -DNA interaction. Actual involvement of the direct repeats in the IolR-DNA interaction was shown by the deficiency of IolR-binding and negative regula tion that was caused by substitution of the conserved bases within the cons erved sequences. These results imply a unique mode of interaction of IolR w ith the target DNA.(C) 1999 Academic Press.