Type VI Secretion System in Pseudomonas aeruginosa

Abstract

Pseudomonas aeruginosa is a Gram-negative bacterium causing chronic infections in cystic fibrosis patients. Such infections are associated with an active type VI secretion system (T6SS), which consists of about 15 conserved components, including the AAA+ ATPase, ClpV. The T6SS secretes two categories of proteins, VgrG and Hcp. Hcp is structurally similar to a phage tail tube component, whereas VgrG proteins show similarity to the puncturing device at the tip of the phage tube. In P. aeruginosa, three T6SSs are known. The expression of H1-T6SS genes is controlled by the RetS sensor. Here, 10 vgrG genes were identified in the PAO1 genome, among which three are co-regulated with H1-T6SS, namely vgrG1a/b/c. Whereas VgrG1a and VgrG1c were secreted in a ClpV1-dependent manner, secretion of VgrG1b was ClpV1-independent. We show that VgrG1a and VgrG1c form multimers, which confirmed the VgrG model predicting trimers similar to the tail spike. We demonstrate that Hcp1 secretion requires either VgrG1a or VgrG1c, which may act independently to puncture the bacterial envelope and give Hcp1 access to the surface. VgrG1b is not required for Hcp1 secretion. Thus, VgrG1b does not require H1-T6SS for secretion nor does H1-T6SS require VgrG1b for its function. Finally, we show that VgrG proteins are required for secretion of a genuine H1-T6SS substrate, Tse3. Our results demonstrate that VgrG proteins are not only secreted components but are essential for secretion of other T6SS substrates. Overall, we emphasize variability in behavior of three P. aeruginosa VgrGs, suggesting that, although very similar, distinct VgrGs achieve specific functions.

Glutathione-Disrupted Biofilms of Clinical Pseudomonas aeruginosa Strains Exhibit an Enhanced Antibiotic Effect and a Novel Biofilm Transcriptome

“…In the PAO1 genome, tle4 and tli4 are upstream of vgrG2a. The coexpression of these genes is in line with previous studies showing that secretion of effector proteins is dependent on the presence of VgrG proteins (52). Our findings are particularly interesting as the T6SS upregulation observed may be a defensive mechanism facilitating survival of disrupted biofilms in environments containing bacterial competitors, such as the polymicrobial CF lung.…”

The Self-Identity Protein IdsD Is Communicated between Cells in Swarming Proteus mirabilis Colonies

“…To confirm the loss of T6SS-mediated transport, IdsA (T6SS_Hcp [PFAM family PF05638]) carrying a C-terminal FLAG epitope tag (N-DYKDDD DK-C) (5) was introduced into CCS05. Export of Hcp homologs, such as IdsA, is a hallmark of T6SS-dependent protein transport and has often been used to evaluate T6SS activity (5,8,9,11,26,43). Furthermore, the export of D, as well as IdsB, is dependent on a functional T6SS and has been correlated with IdsA export (5).…”

A Pseudomonas aeruginosa TIR effector mediates immune evasion by targeting UBAP1 and TLR adaptors

“…The outer membrane fraction was pelleted by ultracentrifugation at 100,000  g for 1 h, washed and resuspended in sonication buffer. The preparation of supernatant samples separation by sodium dodecyl sulphate‐polyacrylamide gel electrophoresis and subsequent immunoblotting has been described previously (Hachani et al , 2011). Immunodetection was conducted using monoclonal antibodies against RNA polymerase (NeoClone) and polyclonal antibodies against PilQ, XcpY (Michel et al , 1998) and LasB.…”