DEAD-box ATPase–marked condensates coordinate compartmentalized translation and antibiotic persistence

Antibiotic-tolerant persisters use dormancy as a bet-hedging strategy to evade lethal antibiotics, undermining therapeutic efficacy. Protein condensates have been implicated in bacterial dormancy, yet how these assemblies orchestrate dormancy entry remains unclear. We evolved persisters that enter dormancy before the stationary phase, most harboring mutations in <i>serS</i>, encoding seryl-transfer RNA synthetase (SerRS). These variants recapitulated persistence induced by serine hydroxamate (SHX), a serine analog and SerRS inhibitor. Both the <i>serS</i> mutation and SHX treatment trigger SerRS sequestration into conserved DEAD-box adenosine triphosphatase-associated condensates, coinciding with growth arrest and dormancy. In vitro, the SerRS variant preferentially partitions into DeaD granules, consistent with its distinct in vivo localization. Microscopy revealed spatially restricted translation silencing within condensates upon SerRS partitioning. Together, these phase-separated condensates act as hubs that coordinate the transition from proliferation to dormancy, paralleling eukaryotic cell fate control via localized translation. Our findings provide mechanistic insight into bacterial persistence and suggest that targeting condensates could help combat antibiotic tolerance and delay resistance.