Regulatory exaptation of the catabolite repression protein (Crp)-cAMP system inPseudomonas putida


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Abstract

SummaryThe genome of the soil bacteriumPseudomonas putidaKT2440 encodes singular orthologues of genescrp(encoding the catabolite repression protein, Crp) andcyaA(adenylate cyclase) ofEscherichia coli.The levels of cAMP formed byP. putidacells were below detection with aDictyosteliumbiosensorin vivo.ThecyaAP.putidagene was transcribedin vivobut failed to complement the lack of maltose consumption of acyaAmutant ofE. coli, thereby indicating thatcyaAP.putidawas poorly translated or rendered non-functional in the heterologous host. Yet, generation of cAMP by CyaAP.putidacould be verified by expressing thecyaAP.putidagene in a hypersensitiveE. colistrain. On the other hand, thecrpP.putidagene restored the metabolic capacities of an equivalentcrpmutant ofE. coli,but not in a doublecrp/cyaAstrain, suggesting that the ability to regulate such functions required cAMP. In order to clarify the breadth of the Crp/cAMP system inP. putida,crpandcyaAmutants were generated and passed through a battery of phenotypic tests for recognition of gross metabolic properties and stress-endurance abilities. These assays revealed that the loss of each gene led in most (but not all) cases to the same phenotypic behaviour, indicating a concerted functionality. Unexpectedly, none of the mutations affected the panel of carbon compounds that can be used byP. putidaas growth substrates, the mutants being impaired only in the use of various dipeptides as N sources. Furthermore, the lack ofcrporcyaAhad little influence on the gross growth fingerprinting of the cells. The poor physiological profile of the Crp-cAMP system ofP. putidawhen compared withE. coliexposes a case of regulatory exaptation, i.e. the process through which a property evolved for a particular function is co-opted for a new use.

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