PFER, AN ENTEROBACTIN-RESPONSIVE ACTIVATOR OF FERRIC ENTEROBACTIN RECEPTOR GENE-EXPRESSION IN PSEUDOMONAS-AERUGINOSA

PfeR (Regulator) and PfeS (Sensor), members of the superfamily of so-c alled two-component regulatory protein pairs, are required for the ent erobactin-inducible production of the ferric enterobactin receptor (Pf eA) in Pseudomonas aeruginosa. A pfeR knockout mutant failed to demons trate enterobactin-inducible expression of a pfeA-lacZ fusion, indicat ing that PfeR acts at the level of pfeA gene expression, Consistent wi th this, PfeR overexpressed in P. aeruginosa bound, in bandshift assay s, the promoter region of pfeA. Such binding was enhanced when PfeR-co ntaining extracts were prepared from cells cultured in the presence of enterobactin, consistent with a model of PfeR as an enterobactin-resp onsive activator of pfeA expression. A region showing homology to the consensus binding sequence for the global iron repressor Fur was ident ified upstream of pfeR, suggesting that the pfeRS operon is iron regul ated. As expected, expression of a pfeR-lacZ fusion in P. aeruginosa w as increased under conditions of iron limitation, Enterobactin failed, however, to provide any enhancement of pfeR-lacZ expression under iro n-limiting conditions, indicating that PfeR does not positively regula te pfeRS expression. A pfeA knockout mutant demonstrated enterobactin- inducible expression of a pfeA-lacZ fusion, indicating that the recept or is not required for the enterobactin inducibility of pfeA gene expr ession, Such mutants show growth, albeit reduced, in enterobactin-supp lemented iron-limiting minimal medium, indicating that a second route of uptake across the outer membrane exists for ferric enterobactin in P. aeruginosa and may be important for the initial induction of pfeA i n response to enterobactin.