Fig. 1: Characterization of Mtb ΔesxBA and Mtb ΔcpsA.
a,c, Mtb esxBA-rv3874-75 locus (a) and cpsA-rv3484 locus (c) in Mtb WT and the respective deletion strains. Black half-arrows depict the primer positions (CmR, chloramphenicol resistance; ZeoR, zeocin resistance; Prom., groEL promoter). b,d, Western blot of EsxA and EsxB from total cell lysates and culture filtrates from Mtb WT, ΔRD1, ΔesxBA, ΔesxBA:BA (n = 3) (b) or Mtb WT, ΔcpsA and ΔcpsA:cpsA strains (n = 1) (d). Ag85 was used as a loading control. e, Growth curves of Mtb WT, ΔesxBA and ΔesxBA:BA. f, Growth curves of Mtb WT, ΔcpsA and ΔcpsA:cpsA strains. g, Thin-layer chromatography analysis of PDIM from Mtb WT, ΔcpsA, ΔcpsA:cpsA, ΔesxBA and ΔesxBA:BA cultures (n = 1). Purified PDIM and extracts from Mtb ΔPDIM were used as controls. h, Quantitative analysis of Mtb WT, ΔesxBA, ΔesxBA:BA (top) and Mtb WT, ΔcpsA, ΔcpsA:cpsA (bottom) area per single cell, 2 h post infection. Representative data of three independent experiments (n = 3 independent wells). Results are shown as mean ± standard error of the mean (s.e.m.). One-way ANOVA followed with Šídák’s multiple comparison test. NS, non-significant. i, Snapshot of live iPSDM infected with Mtb WT, ΔesxBA and ΔesxBA:BA 96 h post infection. Nuclear staining (blue) and Mtb-E2-Crimson (red). Scale bars, 50 µm. j, Quantitative analysis of Mtb replication after infection with Mtb WT, ΔesxBA and ΔesxBA:BA. Mtb area per cell was calculated as fold change, relative to 2 h post infection. Representative data of three independent experiments (n = 3 independent wells). Results are shown as mean ± s.e.m. One-way ANOVA followed with Šídák’s multiple comparison test. ***P < 0.001; NS, non-significant. k, Representative images of fixed iPSDM infected with Mtb WT, ΔcpsA and ΔcpsA:cpsA 96 h post infection. Nuclear staining (blue) and Mtb-E2-Crimson (red). Scale bars, 50 µm. l, Quantitative analysis of Mtb replication after infection with Mtb WT, ΔcpsA and ΔcpsA:cpsA. Mtb area per cell was calculated as fold change, relative to 2 h post infection. Representative data of three independent experiments (n = 3 independent wells). Results are shown as mean ± s.e.m. One-way ANOVA followed with Šídák’s multiple comparisons test **P < 0.002; NS, non-significant.
