Figure 2.

Phage gene 28 is responsible for α subunit cleavage. (A) Overview of the first part of the LUZ19 genome. Genes encoding proteins with predicted function are labeled in green, genes encoding structural proteins are blue. The first ten early genes and the two genes up- and downstream of the viral RNAP (highlighted in light grey) were selected and cloned into a single-copy plasmid, integrated in the P. aeruginosa PAO1 rpoA::strep genome. Dilution series of the cells were plated on LB with 1 mM isopropyl-β-d-thiogalactoside (IPTG) and checked for growth. (B) Pull-down analyses of the α RNAP subunit from cells grown in the presence of the indicated phage proteins (after 30 min induction with 1 mM IPTG). As negative control (NC), P. aeruginosa PAO1 rpoA::strep was used. Gp28 or Rac shows a consistent degradation of the RNAP α subunit. (C) Exponentially growing P. aeruginosa PAO1 cells containing Rac were pulse labeled with ³H-uridine in the presence or absence of 1 mM IPTG. Each data point represents the average of three independent samples with standard deviation. For each timepoint, the scintillation counts for the induced sample were normalized to the non-induced sample at the same timepoint.