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Adenylate cyclase (AC) is the enzyme that catalyzes the transformation of adenosine 5'-triphosphate (ATP) into cyclic adenosine 3’, 5’-monophosphate (cAMP) [AC at UniProtKB].
In Escherichia coli and related bacteria, cAMP controls gene expression [Microbiol Mol Biol Rev]. This control occurs via the Catabolite gene Activator Protein (CAP) [Proc Natl Acad Sci U S A] also called Cyclic AMP Receptor Protein (CRP) [Proc Natl Acad Sci U S A]. CRP exists as a homodimer. Each subunit is composed of two domains connected by a small hinge, the N-terminal cAMP-binding domain and the C-terminal DNA-binding domain [CRP at UniProtKB] [CRP at RCSB PDB] . When bound to cAMP, CRP binds to specific sites upstream of promoters, causing transcriptional activation or repression [Annu Rev Biochem]. Interestingly it has been inferred that CRP optimally 'responds' to physiological cAMP concentrations consequently hindering activation by ligands other than cAMP [J Bacteriol].
The mechanisms of CRP-dependent activation of transcription at class I and II promoters have been described by Busby and Ebright [Medline] and further characterized [Medline] [Biochem Soc Trans] [FEMS Microbiol Lett]. The CRP-cAMP complex can act as a transcriptional regulator by competing for DNA binding or forming complexes with other transcriptional regulators, for example CytR [J Biol Chem] [Medline]. It can also act as a roadblock for RNA polymerase at a class I promoter [J Bacteriol]. Transcriptional repression may result from preventing activation by CRP-cAMP [J Bacteriol]. Also it was suggested that upstream-bound CRP-cAMP can repress transcription by restraining RNA polymerase [mBio].
For structural information visit the 'CAP' page maintained by the Biochemistry Biocomputing Group, University College London. Upon binding of cAMP, the long helices at the dimer interface reposition leading to activation of the CRP-cAMP complex [J Biol Chem]. The mechanism by which CRP-cAMP activates transcription initiation is still under scrutiny [Cell Biochem Funct]. The much-awaited structure of CRP without cAMP was reported in 2009 [Proc Natl Acad Sci U S A] [Kalodimos Group Home Page].
The CRP-cAMP complex is a positive transcriptional regulator of a number of catabolite operons [Proc Natl Acad Sci U S A] and as such plays a role in catabolite repression [Medline]. Catabolite repression, also known as carbon catabolite repression, is the mechanism by which glucose or another rapidly metabolized carbohydrate exerts a continued inhibition on synthesis of catabolic enzymes [Res Microbiol]. Catabolite repression has been extensively studied in enteric bacteria, particularly in the case of the lactose operon [J Bacteriol]. The role of cAMP as an antagonist of catabolite repression [Medline] is well established [Medline]. Cyclic AMP-independent mutants of CRP have been isolated that relieve catabolite repression [Medline].
When catabolite repression occurs, a transcriptome analysis revealed that transcription of specific genes may be enhanced by CRP-cAMP [J Bacteriol]. Groups of genes displaying a transcriptional response under condition of catabolite repression were further defined using transcriptome data [BMC Microbiol]. Also mathematical modeling has been applied to catabolite repression and models were compared for their predictive value [FEBS J].
In 2011 a 'fine-tuning' of transcriptional regulation by CRP-cAMP, via the level of cAMP, was described involving the base-pairing RNA Spot 42 [Mol Cell]. However based on data presented the participation of Spot 42 in catabolite repression may have been overstated.
In addition to its role in regulating transcription of catabolite operons, the CRP-cAMP complex also regulates transcription of many other genes including genes involved in flagella formation [J Bacteriol], biofilm formation [J Bacteriol], nitrogen assimilation [Mol Microbiol] [Nucleic Acids Res], anaerobic growth under nitrate-limited conditions [J Bacteriol], ammonia assimilation [BMC Microbiol], energy metabolism [J Bacteriol], nucleotide biosynthesis [J Bacteriol], iron uptake [FEBS J], membrane functions [J Bacteriol], transport system [J Bacteriol], porin synthesis [Mol Microbiol], outer membrane passage [J Bacteriol], bacterial conjugation [J Bacteriol], osmoprotection [J Bacteriol], stress response [Microbiol Mol Biol Rev], degradation of aromatic compound [Microbiology], small protein synthesis [J Bacteriol], acid resistance [J Bacteriol], stringent response [Cell], cell-to-cell communication [J Bacteriol] [J Bacteriol] [Cell Res], multidrug resistance [J Bacteriol] [Journal Antibiot] [Antimicrob Agents Chemother], cold adaptation [J Bacteriol] and genes encoding enzymes of pharmaceutical importance; for example pga [J Biol Chem]. Furthermore some of the genes that are induced under starvation conditions are subject to transcriptional control by CRP-cAMP [Medline]. In uropathogenic E. coli, CRP-cAMP regulates the formation of pili associated with pyelonephritis (Pap pili) [Mol Microbiol] as well as type 1 fimbriae [PLoS Pathog] [Environ Microbiol]. In pathogenic Salmonella CRP-cAMP regulates genes related to biofilm formation [Microbiology].
Early reports have indicated that CRP-cAMP may regulate the transcription of genes involved in cell division. However, it was later established that CRP-cAMP is not essential for cell division [J Bacteriol].
Even though it was reported that CRP-cAMP regulates the synthesis of enzymes belonging to the tricarboxylic acid (TCA) cycle, the TCA cycle flux does not seem to be affected by the absence of the CRP-cAMP complex [J Bacteriol]. The effect of CRP-cAMP on the TCA cycle flux was however observed under certain specific conditions [J Bacteriol].
Finally, novel predicted binding sites have being identified in Escherichia coli that may be functional for CRP [Proc Natl Acad Sci U S A]. Unknown CRP regulons have also been revealed by using a run-off transcription/microarray analysis (ROMA) [Nucleic Acids Res]. Interestingly, CRP can bind to thousands of different sites without affecting transcription initiation of genes located within these sites [Proc Natl Acad Sci U S A]. Investigation of the effect of a crp deletion indicated that epistasis is a crucial factor in adaptive evolution of Escherichia coli laboratory lines [PLoS Genet].
Escherichia coli cyaA strains lacking adenylate cyclase are viable indicating that cAMP is not essential for survival [J Bacteriol]. However, because of the transcriptional regulation by CRP-cAMP, cyaA strains (or crp strains lacking CRP) present some unique features. For example, they do not grow on a large variety of carbon sources including lactose, maltose, galactose, arabinose, glycerol... [Medline], they are not motile and thereby are incapable of chemotactic responses, they are resistant to antibiotics including fosfomycin [Antimicrob Agents Chemother], fosmidomycin [Biosci Biotechnol Biochem], nalidixic acid, streptomycin and mecillinam [J Bacteriol], they are pH-sensitive for growth [J Bacteriol], and they are better hosts for recombinant expression systems, especially in stationary phase [Medline].
Therefore, by controlling gene expression, cAMP is involved in many aspects of Escherichia coli physiology.
The transcription of the cyaA gene is negatively regulated by CRP-cAMP [J Biol Chem], as well as the transcription of the crp gene [Medline]. However, in both cases the negative transcriptional control by CRP-cAMP does not seem to cause major effects in vivo, at least under current experimental conditions [Medline]. In particular, it was shown by using gene fusion techniques that neither cAMP nor CRP-cAMP plays a major role in transcriptional or translational regulation of cyaA expression [J Bacteriol]. In Salmonella typhimurium, transcription of cyaA is also repressed by CRP-cAMP [Genetics].
The transcription of crp is additionally regulated by the DNA-binding protein FIS, with both FIS and CRP-cAMP being required for repression of crp transcription [EMBO J] [FIS at RCSB PDB]. Incidentally, the study of FIS has led to the concept of a transcription factor acting as a local 'topological homeostat' [Front Biosci]. Additionally in vitro studies have indicated that FIS may play a role in catabolite repression [Mol Microbiol]. A physiological role for FIS in catabolite repression was also reported particularly in a rich growth medium [Microbiology]. Additionally transcription of a plasmid-encoded toxin gene from pathogenic E. coli was reported to be regulated by both FIS and CRP [Mol Microbiol].
The translation of adenylate cyclase mRNA is ineffective [Medline] as to prevent excessive synthesis of adenylate cyclase. This can be attributed to the fact that overproduction of cAMP is lethal to Escherichia coli possibly due to an accumulation of methylglyoxal [J Bacteriol] [Microbiology].
Finally, intracellular cAMP concentrations vary with the carbon source used for bacterial growth. Therefore, any gene whose transcription is regulated by CRP-cAMP may have various levels of expression depending on growth conditions, especially the carbon source available to the bacteria. Correspondingly, the CRP-cAMP complex has been implicated in the establishment of foraging-like behavior when the carbon source is unfavorable [J Biol Chem] [Nucleic Acids Res].
|To Chapter I: In vivo regulation of adenylate cyclase|