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Structural basis for effector transmembrane domain recognition by type VI secretion system chaperones.

ABSTRACT: Type VI secretion systems (T6SSs) deliver antibacterial effector proteins between neighboring bacteria. Many effectors harbor N-terminal transmembrane domains (TMDs) implicated in effector translocation across target cell membranes. However, the distribution of these TMD-containing effectors remains unknown. Here, we discover prePAAR, a conserved motif found in over 6000 putative TMD-containing effectors encoded predominantly by 15 genera of Proteobacteria. Based on differing numbers of TMDs, effectors group into two distinct classes that both require a member of the Eag family of T6SS chaperones for export. Co-crystal structures of class I and class II effector TMD-chaperone complexes from Salmonella Typhimurium and Pseudomonas aeruginosa, respectively, reveals that Eag chaperones mimic transmembrane helical packing to stabilize effector TMDs. In addition to participating in the chaperone-TMD interface, we find that prePAAR residues mediate effector-VgrG spike interactions. Taken together, our findings reveal mechanisms of chaperone-mediated stabilization and secretion of two distinct families of T6SS membrane protein effectors.

SUBMITTER: Ahmad S

PROVIDER: S-EPMC7773334 | biostudies-literature | 2020 Dec

REPOSITORIES: biostudies-literature

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Structural basis for effector transmembrane domain recognition by type VI secretion system chaperones.

Ahmad Shehryar S Tsang Kara K KK Sachar Kartik K Quentin Dennis D Tashin Tahmid M TM Bullen Nathan P NP Raunser Stefan S McArthur Andrew G AG Prehna Gerd G Whitney John C JC

eLife 20201215

Type VI secretion systems (T6SSs) deliver antibacterial effector proteins between neighboring bacteria. Many effectors harbor N-terminal transmembrane domains (TMDs) implicated in effector translocation across target cell membranes. However, the distribution of these TMD-containing effectors remains unknown. Here, we discover prePAAR, a conserved motif found in over 6000 putative TMD-containing effectors encoded predominantly by 15 genera of Proteobacteria. Based on differin ...[more]

PMID: 33320089

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Project description:The type VI secretion system (T6SS) is a bacterial nanoscale weapon that delivers toxins into prey ranging from bacteria and fungi to animal hosts. The cytosolic contractile sheath of the system wraps around stacked hexameric rings of Hcp proteins, which form an inner tube. At the tip of this tube is a puncturing device comprising a trimeric VgrG topped by a monomeric PAAR protein. The number of toxins a single system delivers per firing event remains unknown, since effectors can be loaded on diverse sites of the T6SS apparatus, notably the inner tube and the puncturing device. Each VgrG or PAAR can bind one effector, and additional effector cargoes can be carried in the Hcp ring lumen. While many VgrG- and PAAR-bound toxins have been characterized, to date, very few Hcp-bound effectors are known. Here, we used 3 known Pseudomonas aeruginosa Hcp proteins (Hcp1 to -3), each of which associates with one of the three T6SSs in this organism (H1-T6SS, H2-T6SS, and H3-T6SS), to perform in vivo pulldown assays. We confirmed the known interactions of Hcp1 with Tse1 to -4, further copurified a Hcp1-Tse4 complex, and identified potential novel Hcp1-bound effectors. Moreover, we demonstrated that Hcp2 and Hcp3 can shuttle T6SS cargoes toxic to Escherichia coli. Finally, we used a Tse1-Bla chimera to probe the loading strategy for Hcp passengers and found that while large effectors can be loaded onto Hcp, the formed complex jams the system, abrogating T6SS function. IMPORTANCE The type VI secretion system (T6SS) is an effective weapon used by bacteria to outgrow or kill competitors. It can be used by endogenous commensal microbiota to prevent invasion by pathogens or by pathogens to overcome resident flora and successfully colonize a host or a specific environmental niche. The T6SS is a key contributor to this continuous arms race between organisms as it delivers a multitude of toxins directed at essential processes, such as nucleic acid synthesis and replication, cell wall and membrane integrity, protein synthesis, or cofactor abundance. Many T6SS toxins with unknown function remain to be discovered, whose yet-uncharacterized targets could be exploited for antimicrobial drug design. The systematic search for these toxins is not facilitated by the presence of readily recognizable T6SS motifs, and unbiased screening approaches are thus required. Here, we successfully used a known shuttle for cargo T6SS effectors, Hcp, as bait to identify uncharacterized toxins.

Dataset Information

Structural basis for effector transmembrane domain recognition by type VI secretion system chaperones.

Publications

Structural basis for effector transmembrane domain recognition by type VI secretion system chaperones.

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