The interplay of EIIA(Ntr) with C-source regulation of the Pu promoter of Pseudomonas putida mt-2

The presence of some sugars (e.g. glucose) downregulates the activity of the Pu promoter of plasmid pWW0 of Pseudomonas putida mt-2, which drives the upper TOL operon for biodegradation of m-xylene. Genetic evidence produced 20 years ago documented an effect of the EIIA(Ntr) (PtsN) protein of the nitrogen-related phosphoenolpyruvate-dependent phosphotransferase system (PTSNtr) in such a C-source control of Pu activity. In this study, we have exploited the wealth of recent information on the PTS of P. putida as well as transcriptomic data available in the last few years on this bacterium to revisit this question - and the role of EIIA(Ntr) as such. To this end, we examined Pu output under physiological conditions known to either phosphorylate PTS proteins to saturation or to deplete them altogether from high-energy phosphate. The results showed that Pu activity is checked by EIIA(Ntr) regardless of its phosphorylation state. However, such inhibition is intensified during growth on glucose (which correlates with more phosphate-free EIIA(Ntr)) and partially relieved in fructose, which triggers phosphorylation of PTS proteins. These data explain former inconsistencies on the Pu-PTSNtr interplay and provides a better understanding of the metabolic and regulatory retroactivity between the TOL plasmid and its host metabolism.