The Pseudomonas putida MAD2 strain is derived from P. putida KT2442 and contains a Pu-lacZ fusion and constitutive, inducer-independent xylR variant, allowing experimental focus on the Pu promoter’s IHF dependence. Beta-gal assay of ihfA-variable strains against a growth curve indicated near-total loss of Pu activity in the IHF-mutant strain in every growth phase, while IHF+ strains showed restricted activity in the exponential phase and sharply increased activity in stationary phase. Western blots of the IHF+ strains across the growth phases revealed an approximately seven-fold increase in IHF levels in stationary phase over exponential phase, suggesting Pu dependence on IHF concentration. In vitro IHF binding was visualized by UV footprinting and compared to footprints of rpoN and xylR controls, which indicated IHF binding is independent of both. In vivo UV footprinting of exponential vs. stationary phase ihfA-variable strains correlated to in vitro footprints, and demonstrated IHF binding occurred only in stationary phase.
The paper reports that TF forms complex with other proteins for binding with reported sites.
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|
|TTTTTTACAAAGAAAATCAATAATTTA||xylA, xylM, xylC, xylW, xylU,||