The type VI secretion system (T6SS) is a recently discovered export system that is used by Gram-negative bacteria to deliver effector proteins to other cells in a cell contact-dependent manner. The mechanism of the T6SS and the functions of the secreted effectors are not well understood. Now, Russell et al. show that Tse1 and Tse3, two previously identified substrates of the Pseudomonas aeruginosa T6SS, are effector proteins that degrade the peptidoglycan of other bacteria, conferring a growth advantage on P. aeruginosa.

Tse1 and Tse3 ... are effector proteins that degrade the peptidoglycan of other bacteria, conferring a growth advantage on P. aeruginosa.

Initially, the authors studied the sequences of two T6SS components of unknown function, Tse1 and Tse3, and found that they had motifs that are characteristic of the peptidoglycan-degrading enzymes amidase and muramidase, respectively. Tse1 and Tse3 had peptidoglycan-degrading activity in vitro and caused rapid cell lysis when inducibly expressed and targeted to the periplasm of Escherichia coli. However, E. coli was not lysed when the proteins were cytoplasmically localized, and neither did exogenous Tse1 cause lysis of P. aeruginosa, suggesting that Tse1 and Tse3 require the T6SS to access their substrate in the periplasm of target cells. In competition assays with Pseudomonas putida, which probably inhabits a similar niche to P. aeruginosa, lack of Tse1 and Tse3 reduced P. aeruginosa fitness on solid medium but not in liquid medium, presumably because T6SS function requires cell–cell contact.

As P. aeruginosa can also target its siblings through the T6SS, it must have a mechanism to protect its own peptidoglycan. Russell et al. identified two candidate immunity proteins that are encoded in bicistronic operons with Tse1 and Tse3, naming them Tsi1 and Tsi3. In vitro, Tsi1 and Tsi3 bound to and inactivated Tse1 and Tse3, respectively, and the deletion of tsi1 or tsi3 led to inhibition of P. aeruginosa growth on solid medium. Similarly to the lytic proteins, the authors found that the immunity proteins must be periplasmic to function.

Taken together, these experiments suggest that the T6SS pierces only the outer membrane of the recipient bacterium to allow the effector proteins Tse1 and Tse3 to gain access to the periplasm. The presence of the immunity proteins protects P. aeruginosa from its siblings while allowing it to attack the rival cells around it.