Curation Information

Publication
Roadblock repression of transcription by Bacillus subtilis CodY.;Belitsky BR, Sonenshein AL;Journal of molecular biology 2011 Aug 26; 411(4):729-43 [21699902]
TF
CodY [P39779, view regulon]
Reported TF sp.
Bacillus subtilis subsp. subtilis str. SMY
Reported site sp.
Bacillus subtilis subsp. subtilis str. SMY
Created by
Elliot White
Curation notes
-

Experimental Process

Using a Dnase protection assay, researchers determined the presence of three CodY-protected sites around the transcription start. The one upstream site was seen to only be bound at high concentrations. Researchers then preformed EMSA's and B-gal assays on the two downstream sites, which showed that the sites regulated independantly, and that the first site was tighter bound, and responsible for most of the regulatory effect. Site directed mutagenesis was then used to confirm the role of site 1, and quantify the regulatory effect of site 2.

Transcription Factor Binding Sites


AATATTCTGAAATTT
AAAAGACTGAATATT
AATATTTAAACAATT
AATTATGTGAAAGGT
AATATTCTGAAATTT
AAAAGACTGAATATT
AATATTTAAACAATT
AATTATGTGAAAGGT

Gene Regulation

Regulated genes for each binding site are displayed below. Gene regulation diagrams show binding sites, positively-regulated genes, negatively-regulated genes, both positively and negatively regulated genes, genes with unspecified type of regulation. For each indvidual site, experimental techniques used to determine the site are also given.

Site sequence Regulated genes Gene diagram Experimental techniques TF function TF type
AATATTCTGAAATTT ybgE,
... ... ybgE ybgB ybgA ybgF mmuM glnT ybgJ
Experimental technique details Beta-gal reporter assay - Experimental technique details DNAse footprinting (ECO:0005631) - Experimental technique details EMSA (ECO:0001807) - Experimental technique details Site directed mutagenesis (ECO:0005667) - repressor dimer
AAAAGACTGAATATT ybgE,
... ... ybgE ybgB ybgA ybgF mmuM glnT ybgJ
Experimental technique details Beta-gal reporter assay - Experimental technique details DNAse footprinting (ECO:0005631) - Experimental technique details EMSA (ECO:0001807) - Experimental technique details Site directed mutagenesis (ECO:0005667) - repressor dimer
AATATTTAAACAATT ybgE,
... ... ybgE ybgB ybgA ybgF mmuM glnT ybgJ
Experimental technique details Beta-gal reporter assay - Experimental technique details DNAse footprinting (ECO:0005631) - Experimental technique details EMSA (ECO:0001807) - Experimental technique details Site directed mutagenesis (ECO:0005667) - repressor dimer
AATTATGTGAAAGGT ybgE,
... ... ybgE ybgB ybgA ybgF mmuM glnT ybgJ
Experimental technique details Beta-gal reporter assay - Experimental technique details DNAse footprinting (ECO:0005631) - Experimental technique details EMSA (ECO:0001807) - Experimental technique details Site directed mutagenesis (ECO:0005667) - repressor dimer