Endogenous membrane stress induces T6SS activity inPseudomonas aeruginosa

The type 6 secretion system (T6SS) is a dynamic organelle encoded by many gram-negative bacteria that can be used to kill competing bacterial prey species in densely occupied niches. Some predatory species, such asVibrio cholerae, use their T6SS in an untargeted fashion while in contrast,Pseudomonas aeruginosaassembles and fires its T6SS apparatus only after detecting initial attacks by other bacterial prey cells; this targeted attack strategy has been termed the T6SS tit-for-tat response. Molecules that interact with theP. aeruginosaouter membrane such as polymyxin B can also trigger assembly of T6SS organelles via a signal transduction pathway that involves protein phosphorylation. Recent work suggests that a phospholipase T6SS effector (TseL) ofV. choleraecan induce T6SS dynamic activity inP. aeruginosawhen delivered to or expressed in the periplasmic space of this organism. Here, we report that inhibiting expression of essential genes involved in outer membrane biogenesis can also trigger T6SS activation inP. aeruginosa. Specifically, we developed a CRISPR interference (CRISPRi) system to knock down expression ofbamA,tolB, andlptDand found that these knockdowns activated T6SS activity. This increase in T6SS activity was dependent on the same signal transduction pathway that was previously shown to be required for the tit-for-tat response. We conclude that outer membrane perturbation can be sensed byP. aeruginosato activate the T6SS even when the disruption is generated by aberrant cell envelope biogenesis.