Atomic Structure of T6SS Reveals Interlaced Array Essential to Function

Abstract

Summary Type VI secretion systems (T6SSs) are newly identified contractile nanomachines that translocate effector proteins across bacterial membranes. The Francisella pathogenicity island, required for bacterial phagosome escape, intracellular replication and virulence, was presumed to encode a T6SS-like apparatus. Here, we experimentally confirm the identity of this T6SS and, by cryo electron microscopy (cryoEM), show the structure of its post-contraction sheath at 3.7 Å resolution. We demonstrate the assembly of this T6SS by IglA/IglB and secretion of its putative effector proteins in response to environmental stimuli. The sheath has a quaternary structure with handedness opposite that of contracted sheath of T4 phage tail and is organized in an interlaced two-dimensional array by means of β sheet augmentation. By structure-based mutagenesis, we show that this interlacing is essential to secretion, phagosomal escape, and intracellular replication. Our atomic model of the T6SS will facilitate design of drugs targeting this highly prevalent secretion apparatus.

Cryo‐ EM reconstruction of Type VI secretion system baseplate and sheath distal end

“…Evolutionarily related Type VI secretion system (T6SS) is used by bacteria to deliver proteins into both bacterial and eukaryotic cells (Mougous et al , 2006; Pukatzki et al , 2006; Hood et al , 2010; MacIntyre et al , 2010; Durand et al , 2014; Ho et al , 2014; Alcoforado Diniz et al , 2015; Hachani et al , 2016). The current model of T6SS biogenesis and mode of action is largely based on well‐understood phage assembly (Leiman et al , 2009; Lossi et al , 2011, 2013; Ho et al , 2014; Zoued et al , 2014; Clemens et al , 2015; Kudryashev et al , 2015; Cianfanelli et al , 2016). However, many important features are unknown mostly due to the lack of high‐resolution structural information of T6SS.…”

“…The V. cholerae sheath subunits are interconnected by a mesh of VipA N‐terminal and VipB C‐terminal linkers (Kudryashev et al , 2015) similarly to T6SS sheath of Francisella novicida and R‐type pyocin sheath (Clemens et al , 2015; Ge et al , 2015). The sheath was shown to assemble across the entire width of a cell, and this allowed to use live‐cell fluorescence microscopy to study its dynamics and subcellular localization (Basler et al , 2012; Brunet et al , 2013; Gerc et al , 2015; Zoued et al , 2016; Vettiger et al , 2017).…”

VgrG C terminus confers the type VI effector transport specificity and is required for binding with PAAR and adaptor–effector complex

“…Functional and structural studies have shown that the T6SS nanomachine shares striking similarities with the bacteriophage tail structure (14)(15)(16)(17)(18)(19)(20)(21)(22). Accumulating evidence suggests that the baseplate complex is recruited into the membrane-associated protein complex and initiates the polymerization of a TssB-TssC (VipAVipB) contractile sheath.…”

Francisella tularensistype VI secretion system comes of age

“…Hence it is assumed that IglA/IglB complex is similar to V. cholerae VipA/VipB complex and form tubular transmembrane structure. 21 The intermediate homology exerts DotU and PdpB/Icm and, finally, Francisella's VgrG that is a small 17.5 kDa protein shares only limited homology with the VgrG proteins from V. cholerae. On the other hand Francisella's VgrG protein is also secreted into culture supernatants and into macrophage cytosol.…”