Universal Stress Protein: A Molecular Switch Controlling Stress Survival in Micrococcus luteus

Bacterial dormancy phenotypes are induced by stress and produced stochastically to counteract harsh environmental conditions, including temperature changes, oxidative stress, nutrient deprivation, and antibiotic exposure. Both persistent survival and the viable but nonculturable (VBNC) state phenotypes of dormancy are thought to exist on a continuum, with specific roles in a conglomerate “bet-hedging” strategy. Over 100 species of pathogenic and non-pathogenic bacteria use a dormancy phenotype for survival, including Staphylococcus aureus, Pseudomonas aeruginosa, Escherichia coli, and Mycobacterium tuberculosis. Most human pathogens have antibiotic resistant strains, the production of which have been blamed on tolerance mechanisms in dormancy phenotypes and antimicrobial overuse. Although much work has been done to elucidate the processes involved in resistance, the molecular mechanisms of tolerance are heavily under investigated. Prior proteomic characterization of Micrococcus luteus, a simplified dormancy system, identified UspA616 and 17 other proteins to be upregulated in the dormancy phenotype induced by starvation stress. What role does UspA616 play in the dormancy phenotype of M. luteus? Competition experiments in nutrient deprived acetate minimal media (AMM) (Loebel Model) identified two phenotypes for ΔuspA616::kan knockout; an exponential phase growth advantage and a deficiency in stress fitness during competition with ΔcrtE::kan. These phenotypes were confirmed by fluorescent microscopy as a defect in survival during stress in both the Loebel Model of starvation and the Wayne Hypoxia Model. Proteomic analysis of the ΔuspA616::kan strain of M. luteus by Normalized Spectral Abundance Factor (NSAF) and Accurate Mass Tag (AMT) showed a clear dysregulation of the glyoxylate cycle enzymes isocitrate lyase and malate synthase. With these data the following conclusions were established: (1) M. luteus is a simplified system for studying the molecular mechanisms dictating dormancy phenotypes, (2) UspA616 is critical for stress survival under starvation and hypoxic conditions, matching two models of latency for M. tuberculosis, and (3) UspA616 acts as a survival switch for properly activating and maintaining glyoxylate cycle activity during the response to nutrient starvation. Therefore, UspA616 is a critical protein for the regulation of the dormancy phenotype in M. luteus.