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J. Bacteriol. doi:10.1128/JB.01350-08
Copyright (c) 2008, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved.

Expression of the cpdA Gene Encoding a 3',5'-cyclic AMP (cAMP) Phosphodiesterase Is Positively Regulated by cAMP-CRP Complex

Department of Environmental Science and Protein Research Center for Bio-Industry, Hankuk University of Foreign Studies, Yongin, Kyunggi-Do 449-791; Department of Environmental Medical Biology and Institute of Tropical Medicine, The Brain Korea 21 Project, Yonsei University College of Medicine, Seoul 120-752, South Korea

^* To whom correspondence should be addressed. Email: khlee{at}hufs.ac.kr.

	Abstract

The intracellular level of cyclic 3',5'-adenosine monophosphate (cAMP), a signaling molecule that mediates a variety of cellular processes, is finely modulated by regulation of its synthesis, excretion, and degradation. In this study, cAMP phosphodiesterase (CpdA), an enzyme that catalyzes the conversion of cAMP to AMP, was characterized in a pathogenic bacterium, Vibrio vulnificus. The cpdA gene exists in an operon composed of mutT, yqiB, cpdA, and yqiA, of which transcription was initiated -22 upstream of mutT. A cpdA-null mutant of V. vulnificus contained significantly higher levels of cAMP than the wildtype, but showed no detectable cAMP when a multicopy plasmid of the cpdA gene was provided in trans, suggesting CpdA is responsible for cAMP degradation. Cellular contents of the CpdA protein decreased dramatically in both cya and crp mutants. In addition, expression of the cpdA::luxAB transcription fusion decreased in cya and crp mutants. cpdA::luxAB expression in the cya mutant increased in a concentration-dependent manner upon exogenous addition of cAMP. The cAMP-CRP complex bound directly to the upstream region of mutT, which includes a putative CRP-binding sequence centered at -95.5 relative to the transcription start site. Site-directed mutagenesis or deletion of this sequence in the cpdA::luxAB transcription fusion resulted in loss of regulation by cAMP and CRP. Thus, this study demonstrates that CpdA plays a crucial role in determining the intracellular cAMP level, and shows for the first time that expression of cpdA is activated by the cAMP-CRP complex via direct binding to the regulatory region.