PT  - JOURNAL ARTICLE
AU  - Chiara Rapisarda
AU  - Yassine Cherrak
AU  - Romain Kooger
AU  - Victoria Schmidt
AU  - Riccardo Pellarin
AU  - Laureen Logger
AU  - Eric Cascales
AU  - Martin Pilhofer
AU  - Eric Durand
AU  - Rémi Fronzes
TI  - <em>In situ</em> and high-resolution Cryo-EM structure of the Type VI secretion membrane complex
AID  - 10.1101/441683
DP  - 2018 Jan 01
TA  - bioRxiv
PG  - 441683
4099  - http://biorxiv.org/content/early/2018/10/13/441683.short
4100  - http://biorxiv.org/content/early/2018/10/13/441683.full
AB  - Bacteria have evolved macromolecular machineries that secrete effectors and toxins to survive and thrive in diverse environments. The type VI secretion system (T6SS) is a contractile machine that is related to Myoviridae phages. The T6SS is composed of a baseplate that contains a spike onto which an inner tube is built, surrounded by a contractile sheath. Unlike phages that are released to and act in the extracellular medium, the T6SS is an intracellular machine inserted in the bacterial membranes by a trans-envelope complex. This membrane complex (MC) comprises three proteins: TssJ, TssL and TssM. We previously reported the low-resolution negative stain electron microscopy structure of the enteroaggregative Escherichia coli MC and proposed a rotational 5-fold symmetry with a TssJ:TssL:TssM stoichiometry of 2:2:2. Here, cryo-electron tomography analysis of the T6SS MC confirmed the 5-fold symmetry in situ and identified the regions of the structure that insert into the bacterial membranes. A high resolution model obtained by single particle cryo-electron microscopy reveals its global architecture and highlights new features: five additional copies of TssJ, yielding a TssJ:TssL:TssM stoichiometry of 3:2:2, a 11-residue loop in TssM, protruding inside the lumen of the MC and constituting a functionally important periplasmic gate, and hinge regions. Based on these data, we revisit the model on the mechanism of action of the MC during T6SS assembly and function.