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Identification of CodY targets in Bacillus anthracis by genome-wide in vitro binding analysis
A Château1, W van Schaik, P Joseph
1Unité Toxines et Pathogénie Bactérienne, Département de Microbiologie, Institut Pasteur, Paris, France.
Journal of Bacteriology
|January 8, 2013
Summary
Bacillus anthracis CodY regulates nearly 500 genes, controlling nutrient limitation responses. This study identified direct CodY DNA targets and a consensus binding motif, revealing CodY
Area of Science:
- Microbiology
- Molecular Biology
- Bacterial Genetics
Background:
- CodY is a key transcriptional regulator in Gram-positive bacteria, responding to nutrient availability.
- Bacillus anthracis CodY influences the expression of approximately 500 genes, impacting various cellular processes.
- Understanding CodY's regulatory network is crucial for deciphering bacterial adaptation mechanisms.
Purpose of the Study:
- To identify the direct DNA targets of Bacillus anthracis CodY.
- To elucidate the CodY-binding specificity and consensus motif.
- To analyze the direct regulation of surface layer protein genes (sap and eag) by CodY.
Main Methods:
- Affinity purification of CodY-DNA complexes to identify binding sites.
- Electrophoretic mobility shift assays (EMSA) and DNase I footprinting to confirm binding specificity.
- Analysis of transcriptional lacZ reporter fusions in wild-type and codY mutant strains.
Main Results:
- Identified 389 CodY DNA binding sites, with 132 sites near regulatory regions of 197 genes.
- Determined a Bacillus anthracis CodY-binding consensus motif present in most purified DNA regions.
- Confirmed CodY as a direct repressor of sap and eag gene expression, encoding surface layer proteins.
Conclusions:
- CodY directly binds to specific DNA sequences to regulate gene expression in Bacillus anthracis.
- CodY controls a large regulon, including many genes indirectly, highlighting its broad regulatory role.
- The expression of surface layer proteins is co-regulated by CodY and PagR, integrating metabolic and environmental signals.
