Figure 5.

Employing ectopic expression constructs to study the type IV competence pilus extension ATPase PilB. (A) Natural transformation of the indicated strains was tested. All ectopic expression constructs in these strains were integrated at the VCA0692 locus. IPTG (100 µM), arabinose (0.2%), and/or theophylline (1.5 mM) were added to reactions as indicated below each bar. Data are from at least 3 independent biological replicates and shown as the mean ± SD. Statistical comparisons were made by one-way ANOVA with Tukey’s post-test. LOD, limit of detection, ***P < 0.001. (B) Representative images of surface piliation. All strains harbor ∆pilB and ∆pilT mutations at the native locus and the indicated ectopic pilB expression construct. Where indicated, cells were grown with inducer as follows: 100 µM IPTG for P_tac, 0.2% arabinose for P_BAD, 100 µM IPTG + 1.5 mM theophylline for P_tac-riboswitch, and 0.2% arabinose + 1.5 mM theophylline for P_BAD-riboswitch. Examples of extended pili in no inducer samples are indicated by white arrows. Scale bars, 1 µm.