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Drug-insensitive tumor subpopulations remain a significant barrier to effective cancer treatment. Recent works suggest that within isogenic drug-sensitive cancer populations, subsets of cells can enter a ‘persister’ state allowing them to survive prolonged drug treatment. Such persisters are well-described in antibiotic-treated bacterial populations. In this review, we compare mechanisms of drug persistence in bacteria and cancer. Both bacterial and cancer persisters are associated with slow-growing phenotypes, are metabolically distinct from non-persisters, and depend on the activation of specific regulatory programs. Moreover, evidence suggests that bacterial and cancer persisters are an important reservoir for the emergence of drug-resistant mutants. The emerging parallels between persistence in bacteria and cancer can guide efforts to untangle mechanistic links between growth, metabolism, and cellular regulation, and reveal exploitable therapeutic vulnerabilities.

SummaryHundreds of molecular-level changes within central metabolism allow a cell to adapt to the changing environment. A primary challenge in cell physiology is to identify which of these molecular-level changes are active regulatory events. Here, we introduce pseudo-transition analysis, an approach that uses multiple steady-state observations of 13C-resolved fluxes, metabolites, and transcripts to infer which regulatory events drive metabolic adaptations following environmental transitions. Pseudo-transition analysis recapitulates known biology and identifies an unexpectedly sparse, transition-dependent regulatory landscape: typically a handful of regulatory events drive adaptation between carbon sources, with transcription mainly regulating TCA cycle flux and reactants regulating EMP pathway flux. We verify these observations using time-resolved measurements of the diauxic shift, demonstrating that some dynamic transitions can be approximated as monotonic shifts between steady-state extremes. Overall, we show that pseudo-transition analysis can explore the vast regulatory landscape of dynamic transitions using relatively few steady-state data, thereby guiding time-consuming, hypothesis-driven molecular validations.

Fluxes in microbial metabolism are controlled by various regulatory layers that alter abundance or activity of metabolic enzymes. Recent studies suggest a division of labor between these layers: transcriptional regulation mostly controls the allocation of protein resources, passive flux regulation by enzyme saturation and thermodynamics allows rapid responses at the expense of higher protein cost, and posttranslational regulation is utilized by cells to directly take control of metabolic decisions. We present recent advances in elucidating the role of these regulatory layers, focusing on posttranslational modifications and allosteric interactions. As the systematic mapping of posttranslational regulatory events has now become possible, the next challenge is to identify those regulatory events that are functionally relevant under a given condition.

To counteract oxidative stress and reactive oxygen species (ROS), bacteria evolved various mechanisms, primarily reducing ROS through antioxidant systems that utilize cofactor NADPH. Cells must stabilize NADPH levels by increasing flux through replenishing metabolic pathways like pentose phosphate (PP) pathway. Here, we investigate the mechanism enabling the rapid increase in NADPH supply by exposing Escherichia coli to hydrogen peroxide and quantifying the immediate metabolite dynamics. To systematically infer active regulatory interactions governing this response, we evaluated ensembles of kinetic models of glycolysis and PP pathway, each with different regulation mechanisms. Besides the known inactivation of glyceraldehyde 3-phosphate dehydrogenase by ROS, we reveal the important allosteric inhibition of the first PP pathway enzyme by NADPH. This NADPH feedback inhibition maintains a below maximum-capacity PP pathway flux under non-stress conditions. Relieving this inhibition instantly increases PP pathway flux upon oxidative stress. We demonstrate that reducing cells' capacity to rapidly reroute their flux through the PP pathway increases their oxidative stress sensitivity.