Journal: Nature Communications

Article Title: Dynamic microfluidic single-cell screening identifies pheno-tuning compounds to potentiate tuberculosis therapy

doi: 10.1038/s41467-024-48269-2

Figure Lengend Snippet: a Schematic of the microfluidic dynamic single-cell screening for PTC (colored circles), based on induction of the SOS DNA damage response (light-blue arrow) and decrease in cell-to-cell variation (gradient arrow). The growth and phenotypic transition of rod-shaped bacteria over time ( t ) is also illustrated. The main PTC hit M06 is shown as a blue circle. b Schematic of the expected mode of action and drug potentiation effect of M06. Blunt arrows indicate inhibition and sharp arrows indicate induction. Blue and gray sharp arrows show the entry of M06 into the cell and the exit of its main metabolites, respectively. Bioprocessing of M06 is carried out by unknown nitroreductases (NR) and methyltransferases (MT). Nitroreduction decreases M06 potency and is a likely source of oxidative stress . Reactive oxygen species (ROS) damage lipids, DNA, and may affect NAT (lilac flat arrows). M06 is presumed to inhibit NAT, thus affecting the composition and stability of the mycobacterial cell envelope and impairing the energy metabolism (black flat arrows). NAT might otherwise be implicated in oxidative stress tolerance (orange flat arrow). M06 inhibits DNA gyrase causing DNA breaks. The presence of single-stranded DNA triggers the SOS response. Overall, impairment of cellular energy, lipid, and DNA metabolism might induce additional oxidative stress, which further affects different macromolecules and processes in the cell. Metabolic remodeling and induction of oxidative stress response help the cell countering oxidative damage . This includes the upregulation of the catalase peroxidase KatG, which activates the anti-tubercular drug INH . NAT inhibition by M06 further contributes to INH activation, destabilizing the cell integrity. On the other hand, M06-mediated inhibition of negative DNA supercoiling in the presence of the anti-tubercular drug RIF, which inhibits the RNA polymerase, further impairs DNA topology, preventing both DNA replication and transcription , with lethal consequences for the mycobacterial cell.

Article Snippet: The platform was controlled from the multi-channel Microfluidic Flow Control System (MFCS, Fluigent), via the MAESFLOW Control software (Fluigent): channel (C) 1 drove the FL inlet; C2 drove the CL inlet; C3 drove the reservoirs injecting lids; and C4 drove the FL outlet.

Techniques: Bacteria, Inhibition, Bioprocessing, Activation Assay