Large mutational target size for rapid emergence of bacterial persistence.
Phenotypic heterogeneity displayed by a clonal bacterial population permits a small fraction of cells to survive prolonged exposure to antibiotics. Although first described over 60 y ago, the molecular mechanisms underlying this behavior, termed persistence, remain largely unknown. To systematically explore the genetic basis of persistence, we selected a library of transposon-mutagenized Escherichia coli cells for survival to multiple rounds of lethal ampicillin exposure. Application of microarray-based genetic footprinting revealed a large number of loci that drastically elevate persistence frequency through null mutations and domain disruptions. In one case, the C-terminal disruption of methionyl-tRNA synthetase (MetG) results in a 10,000-fold higher persistence frequency than wild type. We discovered a mechanism by which null mutations in transketolase A (tktA) and glycerol-3-phosphate (G3P) dehydrogenase (glpD) increase persistence through metabolic flux alterations that increase intracellular levels of the growth-inhibitory metabolite methylglyoxal. Systematic double-mutant analyses revealed the genetic network context in which such persistent mutants function. Our findings reveal a large mutational target size for increasing persistence frequency, which has fundamental implications for the emergence of antibiotic tolerance in the clinical setting.
Department of Molecular Biology, Princeton University, Princeton, NJ 08544, USA.
Hany S
HS
Girgis
Kendra
K
Harris
Saeed
S
Tavazoie
D000667:Ampicillin / Q000494:pharmacology*; D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D018983:DNA Footprinting; D024881:Drug Resistance, Bacterial / Q000187:drug effects / Q000235:genetics; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism; D009154:Mutation; D020411:Oligonucleotide Array Sequence Analysis
D000900:Anti-Bacterial Agents; D029968:Escherichia coli Proteins; D000667:Ampicillin