The Type VI Secretion TssEFGK-VgrG Phage-Like Baseplate Is Recruited to the TssJLM Membrane Complex via Multiple Contacts and Serves As Assembly Platform for Tail Tube/Sheath Polymerization

PLoS Genet. 2015 Oct 13;11(10):e1005545. doi: 10.1371/journal.pgen.1005545. eCollection 2015 Oct.

Abstract

The Type VI secretion system (T6SS) is a widespread weapon dedicated to the delivery of toxin proteins into eukaryotic and prokaryotic cells. The 13 T6SS subunits assemble a cytoplasmic contractile structure anchored to the cell envelope by a membrane-spanning complex. This structure is evolutionarily, structurally and functionally related to the tail of contractile bacteriophages. In bacteriophages, the tail assembles onto a protein complex, referred to as the baseplate, that not only serves as a platform during assembly of the tube and sheath, but also triggers the contraction of the sheath. Although progress has been made in understanding T6SS assembly and function, the composition of the T6SS baseplate remains mostly unknown. Here, we report that six T6SS proteins-TssA, TssE, TssF, TssG, TssK and VgrG-are required for proper assembly of the T6SS tail tube, and a complex between VgrG, TssE,-F and-G could be isolated. In addition, we demonstrate that TssF and TssG share limited sequence homologies with known phage components, and we report the interaction network between these subunits and other baseplate and tail components. In agreement with the baseplate being the assembly platform for the tail, fluorescence microscopy analyses of functional GFP-TssF and TssK-GFP fusion proteins show that these proteins assemble stable and static clusters on which the sheath polymerizes. Finally, we show that recruitment of the baseplate to the apparatus requires initial positioning of the membrane complex and contacts between TssG and the inner membrane TssM protein.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Bacteriophages / genetics
  • Bacteriophages / ultrastructure
  • Contractile Proteins / genetics
  • Contractile Proteins / ultrastructure*
  • Cytoplasm / genetics
  • Escherichia coli / genetics
  • Escherichia coli / ultrastructure
  • Escherichia coli Proteins / genetics*
  • Membrane Proteins / genetics
  • Microscopy, Fluorescence
  • Multiprotein Complexes / genetics
  • Sequence Homology, Amino Acid
  • Type VI Secretion Systems / genetics
  • Type VI Secretion Systems / ultrastructure*
  • Viral Proteins / genetics
  • Viral Tail Proteins / genetics*

Substances

  • Contractile Proteins
  • Escherichia coli Proteins
  • Membrane Proteins
  • Multiprotein Complexes
  • Type VI Secretion Systems
  • Viral Proteins
  • Viral Tail Proteins
  • gene 5 protein, Enterobacteria phage T4

Grants and funding

This work was supported by the Centre National de la Recherche Scientifique, the Aix-Marseille Université and grants from the Agence Nationale de la Recherche (ANR-10-JCJC-1303-03 and ANR-14-CE14-0006-02) to EC. YRB and AZ were supported by doctoral fellowships from the French Ministry of Research. AZ was supported by a fellowship from the Fondation pour la Recherche Médicale (FDT20140931060). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.