Bacillus anthracis TIR Domain-Containing Protein Localises to Cellular Microtubule Structures and Induces Autophagy.

Emil Carlsson,Helen Flick-Smith,Dominic C. Jenner,Abigail M. Spear,Helen S. Atkins,Joanne E. Thwaite,Jeak Ling Ding,Bernadette Byrne,Nick Gay

doi:10.1371/journal.pone.0158575

Abstract

Toll-like receptors (TLRs) recognise invading pathogens and mediate downstream immune signalling via Toll/IL-1 receptor (TIR) domains. TIR domain proteins (Tdps) have been identified in multiple pathogenic bacteria and have recently been implicated as negative regulators of host innate immune activation. A Tdp has been identified in Bacillus anthracis, the causative agent of anthrax. Here we present the first study of this protein, designated BaTdp. Recombinantly expressed and purified BaTdp TIR domain interacted with several human TIR domains, including that of the key TLR adaptor MyD88, although BaTdp expression in cultured HEK293 cells had no effect on TLR4- or TLR2- mediated immune activation. During expression in mammalian cells, BaTdp localised to microtubular networks and caused an increase in lipidated cytosolic microtubule-associated protein 1A/1B-light chain 3 (LC3), indicative of autophagosome formation. In vivo intra-nasal infection experiments in mice showed that a BaTdp knockout strain colonised host tissue faster with higher bacterial load within 4 days post-infection compared to the wild type B. anthracis. Taken together, these findings indicate that BaTdp does not play an immune suppressive role, but rather, its absence increases virulence. BaTdp present in wild type B. anthracis plausibly interact with the infected host cell, which undergoes autophagy in self-defence.

Highlights

Toll-like receptors (TLRs) are a family of type I integral membrane proteins with key roles in the innate immune response, the first line of defence against invading pathogens [1]
The BaTdp sequence contains the Box 1 and Box 2 regions characteristic of the Toll/IL-1 receptor (TIR) domains, the 3D structure prediction tool, FUGUE [34], did not find any highly scoring matches. This suggests that at the level of its primary sequence, part of the BaTdp is homologous to TIR domains but the protein may differ at a tertiary level from previously solved structures of TIR domain proteins
This paper details an investigation into the TIR-containing protein from the highly pathogenic bacterial species, B. anthracis, and its overall effect on virulence

Summary

Introduction

Toll-like receptors (TLRs) are a family of type I integral membrane proteins with key roles in the innate immune response, the first line of defence against invading pathogens [1]. The TLRs contain an extracellular domain comprised of a leucine-rich repeat linked via a single transmembrane region to an intracellular Toll/interleukin 1 receptor (TIR). Central to both the initiation and propagation of TLR signalling are heterotypic TIR-TIR interactions involving the TLRs and cytosolic adaptor proteins. MAL and TRAM are bridging adaptors mediating recruitment of MyD88 and TRIF, respectively, to active TLRs, both MyD88 and TRIF can interact directly with some TLRs [3] This in turn is thought to cause association of other proteins crucial in TLR signalling, into a multi-protein complex called a Supramolecular Organizing Centre (SMOC) [4]. SARM has been shown to associate with cytoskeletal structures [8] and regulate microtubule stability via tubulin acetylation [9]

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