DesT was determined to bind to the desCB operon promoter region of P. aeruginosa via EMSA; observation of two DNA-protein complexes indicated two binding sites. An in-silico palindrome search of the DesT-desCB intergenic region identified two putative binding sites (P1, P3), while visual inspection identified a third (P2). Further EMSAs used oligonucleotide probes of all three sites, which demonstrated binding in two of the three (P1 and P2, but not P3). This binding was confirmed by DNase I footprinting and a second EMSA using WT DNA (WT, P1-, P1-/P2-) rather than synthesized probes. Further EMSA was performed in the presence of saturated or unsaturated acyl-CoA, which affected binding of DesT to P2 but not to P1. qPCR of DesT variable strains grown in saturated or unsaturated acyl-CoA media further indicated that P1 had little to no effect on desCB regulation, and that regulation arose primarily from DesT-P2 binding.
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|