Systematic dissection of the agrobacterium type VI secretion system reveals machinery and secreted components for subcomplex formation

PLoS One. 2013 Jul 5;8(7):e67647. doi: 10.1371/journal.pone.0067647. Print 2013.


The type VI secretion system (T6SS) is widely distributed in pathogenic Proteobacteria. Sequence and structural analysis of T6SS reveals a resemblance to the T4 bacteriophage tail, in which an outer sheath structure contracts an internal tube for injecting nucleic acid into bacterial cells. However, the molecular details of how this phage tail-like T6SS structure is assembled in vivo and executed for exoprotein or effector secretion remain largely unknown. Here, we used a systematic approach to identify T6SS machinery and secreted components and investigate the interaction among the putative sheath and tube components of Agrobacterium tumefaciens. We showed that 14 T6SS components play essential roles in the secretion of the T6SS hallmark exoprotein Hcp. In addition, we discovered a novel T6SS exoprotein, Atu4347, that is dispensable for Hcp secretion. Interestingly, Atu4347 and the putative tube components, Hcp and VgrG, are mainly localized in the cytoplasm but also detected on the bacterial surface. Atu4342 (TssB) and Atu4341 (TssC41) interact with and stabilize each other, which suggests that they are functional orthologs of the sheath components TssB (VipA) and TssC (VipB), respectively. Importantly, TssB interacts directly with the three exoproteins (Hcp, VgrG, and Atu4347), in which Hcp also interacts directly with VgrG-1 on co-purification from Escherichia coli. Further co-immunoprecipitation and pulldown assays revealed these subcomplex(es) in A. tumefaciens and thereby support T6SS functioning as a contractile phage tail-like structure.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Agrobacterium / genetics
  • Agrobacterium / metabolism*
  • Bacterial Proteins / genetics
  • Bacterial Proteins / metabolism*
  • Bacterial Secretion Systems / physiology*
  • Cell Membrane / metabolism
  • Gene Order
  • Multigene Family
  • Multiprotein Complexes / metabolism
  • Mutagenesis
  • Operon
  • Promoter Regions, Genetic
  • Protein Binding
  • Protein Stability
  • Transcription, Genetic


  • Bacterial Proteins
  • Bacterial Secretion Systems
  • Multiprotein Complexes

Grant support

This work was in part supported by a National Science Council grant (NSC 98-2311-B-001 -002 -MY3) and a grant from Academia Sinica to EML. JSL and LSM were in part supported by Academia Sinica Summit project. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.