Fig. 5: Rv3706c and Rv3705A mitigate redox stress to permit Mtb persistence.

a Mtb intracellular survival in activated murine BMDMs is attenuated upon CRISPRi-mediated silencing of Rv3706c or Rv3705A. VICE strains were pre-induced with 200 ng/mL ATc for 72 h prior to infection, and ATc was maintained in the culture medium for the duration of the infection. Each time point represents the mean recovered CFU of two biological replicates each performed in triplicate, with error bars denoting SD. Statistical analysis of VICE strains with or without ATc was performed using an unpaired two-tailed t-test at 72 h. b Principal component analysis (PCA) of the Mtb proteome upon CRISPRi-mediated gene silencing of Rv3706c and Rv3705A reveals significant remodelling of physiology. c Linear regression analysis of the dysregulated Mtb proteome reveals significant overlap upon Rv3706c and Rv3705A gene silencing. Each point represents a significantly differentially abundant protein (adj. P < 10⁻⁵) in each respective comparison (genotype ± ATc). Line of best fit is shown in each comparison (pH 7 and pH 5.4). In each comparison, P < 0.001. d Genetic silencing of Rv3706c and Rv3705A alters several biological pathways including redox homoeostasis in Mtb. Heatmap comparing differential protein abundance as log₂(fold-change) of each VICE strain, and a control VICE strain (Ctrl), calculated from three biological replicates. All proteins are below the statistical significance cut-off (adj. P < 10⁻⁵). Complete protein lists are supplied in Supplementary Data 2 and 3. e Depletion of Rv3706c or Rv3705A in Mtb promotes the accumulation of endogenous reactive oxygen species (ROS). VICE strains were grown in 7H9 broth at the indicated pH in the presence or absence of ATc and then stained with the fluorescent dye CellROX Red for 2 h. Each bar represents the median fluorescence intensity (MFI) from three independent cultures (open circles). Statistical analysis was performed using an unpaired two-tailed t-test. Error bars denote SD. f CRISPRi-mediated silencing of Rv3706c or Rv3705A enhances Mtb sensitivity to oxidative stress at acidic pH. Each bar represents mean survival of each VICE strain from three independent cultures (open circle), relative to an unchallenged control assayed in parallel. Error bars denote SD. Statistical analysis was performed using an unpaired two-tailed t-test. Source data are provided as a Source Data file.