Figure 8.

The bioinformatically predicted Mcr11 base‐pairing sequence is required for the repression of lipB expression in Mtb. (a) The organization of the dlaT‐lipB locus, with the internal promoter:GFPv fusion reporter construct and the disrupted potential base‐pairing interactions between the lipB mRNA and Mcr11 is shown below. The bases in the lipB promoter that were mutagenized are indicated in underlined italics. (b) Mtb was grown for 14 days in under hypoxic (1.3% O₂, 5% CO₂), shaking conditions in −OA media (7H9 + 0.2% glycerol, 10% ADC and 0.05% Tyloxapol). Expression from the lipB internal promoter was measured using a GFPv fluorescence assay used to measure promoter activity at the indicated time points. Fluorescence is normalized to the OD₆₂₀ of each sample. Expression from the lipB promoter with wild‐type sequence is shown in black bars and expression from the mutagenized promoter is shown in slashed bars. Results are the means of three biological replicates. Statistical analysis conducted with a two‐tailed Student’s T‐test to compare the expression of the lipB promoter with wild‐type sequence to the mutagenized promoter at each time point. Asterisks indicate significance as follows: *p < .05, **p < .01, ***p < .001, ****p < .0001 [Colour figure can be viewed at https://www.wileyonlinelibrary.com]