growing-mycobacterial-biofilm-as-model-to-study-antimicrobial

Growing Mycobacterial Biofilm as a Model to Study Antimicrobial Resistance

Many bacteria thrive in intricate natural communities, exhibiting key attributes of multicellularity such as communication, cooperation, and competition. ...

Tata Institute of Fundamental Research

The bacterial respiratory electron transport system (ETS) is branched to allow condition-specific modulation of energy metabolism. There is a detailed understanding of the structural and biochemical features of respiratory enzymes; however, a holistic examination of the system and its plasticity is lacking. Here we generate four strains of Escherichia coli harboring unbranched ETS that pump 1, 2, 3, or 4 proton(s) per electron and characterized them using a combination of synergistic methods (adaptive laboratory evolution, multi-omic analyses, and computation of proteome allocation). We report that: (a) all four ETS variants evolve to a similar optimized growth rate, and (b) the laboratory evolutions generate specific rewiring of major energy-generating pathways, coupled to the ETS, to optimize ATP production capability. We thus define an Aero-Type System (ATS), which is a generalization of the aerobic bioenergetics and is a metabolic systems biology description of respiration and its inherent plasticity.

Laboratory evolution of synthetic electron transport system variants reveals a larger metabolic respiratory system and its plasticity.

Relationships between the genome, transcriptome, and metabolome underlie all evolved phenotypes. However, it has proved difficult to elucidate these relationships because of the high number of variables measured. A recently developed data analytic method for characterizing the transcriptome can simplify interpretation by grouping genes into independently modulated sets (iModulons). Here, we demonstrate how iModulons reveal deep understanding of the effects of causal mutations and metabolic rewiring. We use adaptive laboratory evolution to generate E. coli strains that tolerate high levels of the redox cycling compound paraquat, which produces reactive oxygen species (ROS). We combine resequencing, iModulons, and metabolic models to elucidate six interacting stress-tolerance mechanisms: (1) modification of transport, (2) activation of ROS stress responses, (3) use of ROS-sensitive iron regulation, (4) motility, (5) broad transcriptional reallocation toward growth, and (6) metabolic rewiring to decrease NADH production. This work thus demonstrates the power of iModulon knowledge mapping for evolution analysis.

Laboratory evolution, transcriptomics, and modeling reveal mechanisms of paraquat tolerance.

Energy generation pathways are a potential avenue for the development of novel antibiotics. However, bacteria possess remarkable resilience due to the compensatory pathways, which presents a challenge in this direction. NADH, the primary reducing equivalent, can transfer electrons to two distinct types of NADH dehydrogenases. Type I NADH dehydrogenase is an enzyme complex comprising multiple subunits and can generate proton motive force (PMF). Type II NADH dehydrogenase does not pump protons but plays a crucial role in maintaining the turnover of NAD+. To study the adaptive rewiring of energy metabolism, we evolved an  mutant lacking type II NADH dehydrogenase. We discovered that by modifying the flux through the tricarboxylic acid (TCA) cycle,  could mitigate the growth impairment observed in the absence of type II NADH dehydrogenase. This research provides valuable insights into the intricate mechanisms employed by bacteria to compensate for disruptions in energy metabolism.

Article	Year
Laboratory evolution of synthetic electron transport system variants reveals a larger metabolic respiratory system and its plasticity. Nature communications\| PubMed ID: 35760776	2022
Laboratory evolution, transcriptomics, and modeling reveal mechanisms of paraquat tolerance. Cell reports\| PubMed ID: 37713311	2023
TCA cycle tailoring facilitates optimal growth of proton-pumping NADH dehydrogenase-dependent . Microbiology spectrum\| PubMed ID: 37855642	2023

Amitesh Anand

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