Activation of the Cryptic PhnE Permease Promotes Rapid Adaptive Evolution in a Population of Escherichia coli K-12 Starved for Phosphate

Escherichia coli K-12 suffers acetic acid stress during prolonged incubation in glucose minimal medium containing a limiting concentration of inorganic phosphate (0.1 mM Pi ), which decreases the number of viable cells from 6 × 108 to ≤10 CFU/ml between days 6 and 14 of incubation. Here we show that following two serial transfers into Pi -limiting medium, evolved mutants survived prolonged incubation (≈107 CFU/ml on day 14 of incubation). The evolved strains that overtook the populations were generally PhnE+ , whereas the ancestral K-12 strain carries an inactivephnE allele, which prevents the transport of phosphonates. The switching inphnE occurred with a high frequency as a result of the deletion of an 8-bp repeated sequence. In a mixed culture starved for Pi that contained the K-12 ancestral strain in majority, evolved strains grew through PhnE-dependent scavenging of probably organic phosphate esters (not phosphonates or Pi ) released byE. coli K-12 between days 1 and 3, before acetic acid excreted byE. coli K-12 reached toxic levels. The growth yield ofphnE + strains in mixed culture was dramatically enhanced by mutations that affect glucose metabolism, such as anrpoS mutation inactivating the alternative sigma factor RpoS. The long-term viability of evolved populations was generally higher when the ancestral strain carried an inactive rather than an activephnE allele, which indicates that cross-feeding of phosphorylated products as a result of thephnE polymorphism may be essential for the spread of mutants which eventually help populations to survive under Pi starvation conditions.