The astonishing proposal by K. Kimata and coworkers, that cAMP does not play a key role in the exemplary glucose-lactose diauxie, was published in 1997 [Proc Natl Acad Sci U S A]. This is contrary to the 'dogma' that full manifestation of diauxie requires both inducer exclusion, i.e., the capability of glucose to prevent the entry of lactose, and a low level of cAMP.
The authors proposed that inducer exclusion was solely responsible for the glucose-lactose diauxie or, as stated, that "the glucose-lactose diauxie is not mediated by a decreased concentration of CRP-cAMP". The role of CRP-cAMP was reported to be 'crucial' only for activating the transcription of ptsG (encoding the glucose permease).
K. Kimata and coworkers however did not address specific questions. Why is a typical diauxic lag eliminated by addition of cAMP? Why is constitutive β-galactosidase synthesis repressed by glucose? Why is differential rate of β-galactosidase synthesis in fully-induced cells growing exponentially on glucose less than that in cells growing exponentially on glycerol or succinate?
And finally, why mannitol and not fructose [Microbiol Mol Biol Rev] or mannose or sorbitol, can substitute for glucose in producing diauxie?
Indubitably, answering these questions will trigger reconsideration of the proposal that cAMP is not a key element in diauxie.
First posted: Friday, September 5, 2003
The phosphorylation state of Enzyme IIAGlc is essential to adenylate cyclase activity. However, is the statement by B. Hogema and coworkers [Mol Microbiol] that "transport and phosphorylation of PTS carbohydrates cause dephosphorylation of Enzyme IIAGlc" valid? If this dephosphorylation is well documented in the case of the PTS-sugar glucose, it remains to be established for other PTS-sugars.
Transport of the PTS-sugar fructose, for example, does not seem to cause excessive dephosphorylation of Enzyme IIAGlc, as shown by the relatively high level of cAMP measured in fructose-grown wild type strains. Only in fruR strains (lacking the fructose repressor) fructose transport is causing Enzyme IIAGlc to be unphosphorylated, due to competition for phosphorylation [Reviewed in Escherichia coli adenylate cyclase homepage, Chapter III].
However, the above statement is likely to be true. Dephosphorylation of Enzyme I and possibly HPr by transport and concomitant phosphorylation of PTS-sugars other than glucose, may limit phosphorylation of Enzyme IIAGlc, depending on the rate of phosphate transfer.
B. Hogema and coworkers have also reported that, in the absence of PTS transport, the [PEP]/[pyruvate] ratio determines the phosphorylation state of Enzyme IIAGlc [Mol Microbiol].
It was originally proposed by N. Weigel and coworkers that the [PEP]/[pyruvate] ratio plays a role in regulating the PTS, the ratio of [Enzyme I]/[phosphorylated Enzyme I] being determined by the [PEP]/[pyruvate] ratio [J Biol Chem]. Such proposal was substantiated by studies showing that PEP promotes dimerization of Enzyme I while pyruvate triggers the opposite effect [Protein Sci]. The PEP to pyruvate ratio could therefore be a factor in determining adenylate cyclase activity by establishing the phosphorylation state of Enzyme I and therefore indirectly of Enzyme IIAGlc.
In the absense of PTS transport, the correlation between the [PEP]/[pyruvate] ratio and the phosphorylation state of Enzyme IIAGlc remains to be clearly established, especially in growing cells. Among the three carbon sources tested by B. Hogema and coworkers, one (gluconate) did not seem to trigger the expected correlation [Mol Microbiol, see figure 7, panel A, B and C]. Furthermore, the wild type strain used in their studies (MG1655 from the E. coli Genetic Stock Center) present some features that may not be suitable to establish the present correlation [J Bacteriol].
Posted: Saturday, September 24, 2005
In a 2005 article, A. J. Schmidt and coworkers clearly demonstrated that the phosphodiesterase activity of Escherichia coli Dos (Direct oxygen sensor) is c-di-GMP specific [J Bacteriol]. Contrary to what was previously reported [J Biol Chem], they also showed that Dos does not possess a cAMP phosphodiesterase activity. Furthermore the activity of similar phosphodiesterase is actually quite specific for c-di-GMP [J Appl Physiol]. In this context, the most recent proposal [JB, October 2005] that Dos plays an important role in regulating the cAMP level in E. coli under aerobic conditions [J Bacteriol] is surprising and needs to be evaluated.
The proposal that Dos directly regulates the cAMP level in E. coli is based on cAMP measurements in dos mutant strains characterized by the authors as follows. When grown in aerobic conditions these strains suffered serious growth defect, particularly cell division was impaired, as shown by lack of septation resulting in the formation of filaments. Does the large increase of cAMP observed in dos strains, as compared to wild type strains, correlate with their morphology -as inferred by the authors?
Cyclic AMP does not play a significant role in cell division, at least in aerobic conditions as previously demonstrated [J Bacteriol]. To justify the correlation between cAMP level and strain morphology the authors made reference to the effect of cAMP on fic (filamentation induced by cAMP) mutant strains at elevated temperature. However such effect is specifically dependent on both cAMP and a temperature-sensitive factor [J Bacteriol]. A more likely explanation is the excess of cAMP in dos strains may correlate with the observed growth defect due to toxic production of methylglyoxal. Previous studies have shown that an increase in exogenous cAMP causes an increase in methylglyoxal under specific conditions [J Bacteriol]. Conversely, the production of filaments due to impaired cell division has not been linked to methylglyoxal production, which is usually observed in conditions where the methylglyoxal bypass is activated. The authors also reported that in strains overproducing Dos the concentration of cAMP was reduced, as compared to wild type strains, and formation of minicells was observed. Genetically-characterized minicell-producing mutants do not require cAMP for minicells formation [J Bacteriol] thus indicating that a decrease in cAMP concentration is unlikely to trigger the formation of minicells. Ergo, the proposed contention by the authors that a correlation between cAMP concentration and strain morphology is indicative of cAMP being a substrate for Dos in vivo is quite questionable.
The same proposal relied on cAMP measurements performed with strains entering stationary phase and therefore [Mol Microbiol] submitted to multiple stress signals. Unfortunately the molecular mechanisms regulating the cAMP levels during entry into stationary phase are not known. In any case, it is beyond conclusion that Dos plays an important role in regulating the cyclic AMP level in exponentially growing wild type E. coli strains -as suggested by the title of the JB article.
In an opposite frame of thought the authors suggested the possibility that cAMP levels in E. coli may not be directly regulated by the phosphodiesterase activity of Dos. Surprisingly, they proposed that E. coli cAMP phosphodiesterase (CpdA) may be active in reducing the cAMP level in wild type strains, suggesting a role for CpdA in regulating the cAMP levels during entry into stationary phase. It is surely surprising considering the sharp increase in cAMP observed during entry in stationary phase occurs both in E. coli wild type and mutant strains defective in their cAMP phosphodiesterase [J Bacteriol].
What triggers the growth defect of dos mutant strains remains a mystery. Is it the absence of the c-di-GMP phosphodiesterase activity of Dos or the lack of signal transduction by the heme-binding domain of Dos? These questions may be answered when the role of Dos in E. coli is finally discovered.
Note: A cAMP phosphodiesterase activity for Dos (DosP) was again reported in 2014 based on faulty rational [Biotechnol Bioeng]