Figure 1. Bacteriophage HK022 integrates between the signaling genes torS and torT, disrupting regulation of torCAD and a metabolic bet-hedging strategy.
(A) HK022 integrates as a prophage at an integration site (attBHK022) between torS and torT, separating torS from the IscR binding site that represses its transcription. TorS regulates torCAD by phosphorylating and dephosphorylating the transcription factor TorR, which in its phosphorylated state activates transcription from the torCAD promoter. To phosphorylate TorR, TorS must interact with TMAO-bound TorT; in the absence of this interaction, TorS dephosphorylates TorR. When oxygen is present, transcription of torS and torT is repressed to an extremely low level by IscR, and stochasticity in the ratio of TorS to TorT leads to noisy torCAD transcription (Carey et al., 2018). (B) The HK022 prophage shuts off aerobic transcription of torCAD but leaves anaerobic expression intact. Distributions of single-cell fluorescence are shown for strains carrying a fluorescent reporter of torCAD transcription. Data are shown for an HK022 lysogen (DFE12) and a non-lysogen (MMR8) grown in the presence or absence of oxygen. Each circle represents a fluorescence measurement made in an individual cell. To facilitate qualitative comparisons between distributions, density curves (shown in gray) were generated from single-cell measurements (see Materials and methods). Data are pooled from three independent experiments, with the vertical red lines indicating the population mean fluorescence for each experiment. a.u., arbitrary units. (C,D) Most cells carrying the HK022 prophage fail to grow following rapid oxygen depletion. Each circle represents an individual cell monitored for growth following an aerobic-to-anaerobic transition. The same data are presented on a linear scale (C) for easier comparison with (B) and on a log scale (D) for clearer resolution of individual points. The HK022 lysogen (JNC173) constitutively expresses CFP to distinguish it from the non-lysogen (JNC174), which constitutively expresses mCherry. Both strains carry the YFP reporter of torCAD transcription and lack fhuA, the gene encoding the HK022 receptor. Growth is quantified as the ratio of microcolony area approximately 5 hr after oxygen depletion to the area of the parent cell at the time of depletion. Data are shown for a single representative experiment.