FIG 2.

AtxA and CO₂/bicarbonate-dependent transcriptional activation of lef and cya promoters correlated with direct binding of AtxA to the promoters. (A) GFP fluorescence levels due to transcriptional activation of the pagA, cya, and lef promoters in pRSP, Prpe, and pRPL plasmids, respectively. High fluorescence was seen only in AtxA-expressing strains (+pKA2 plasmid) grown in CO₂/bicarbonate containing medium. Differences were analyzed by Welch’s t test. (B) EMSA with infrared dye-labeled fragments of the cya and lef promoters and the indicated concentrations of AtxA protein. Samples were run on 2% agarose in EABE buffer. (C) Competitive EMSA with 750 nM AtxA and labeled cya and lef promoter fragments (1.7 nM) mixed with unlabeled promoter fragments (competitor probes) in 10×, 60×, and 250× molar ratios. Arrows indicate shifted fragments.