A Multifunctional Region of the Shigella Type 3 Effector IpgB1 Is Important for Secretion from Bacteria and Membrane Targeting in Eukaryotic Cells

Sonia C P Costa,Cammie F Lesser,Daniel E Voth

doi:10.1371/journal.pone.0093461

Sonia C P Costa, Cammie F Lesser + Show 1 more

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https://doi.org/10.1371/journal.pone.0093461

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Journal: PloS one	Publication Date: Apr 9, 2014
Citations: 37	License type: CC BY 4.0

Affiliation: Massachusetts General Hospital, Harvard University

Abstract

Type 3 secretion systems are complex nanomachines used by many Gram–negative bacteria to deliver tens of proteins (effectors) directly into host cells. Once delivered into host cells, effectors often target to specific cellular loci where they usurp host cell processes to their advantage. Here, using the yeast model system, we identify the membrane localization domain (MLD) of IpgB1, a stretch of 20 amino acids enriched for hydrophobic residues essential for the targeting of this effector to the plasma membrane. Embedded within these residues are ten that define the IpgB1 chaperone-binding domain for Spa15. As observed with dedicated class IA chaperones that mask hydrophobic MLDs, Spa15, a class IB chaperone, promotes IpgB1 stability by binding this hydrophobic region. However, despite being stable, an IpgB1 allele that lacks the MLD is not recognized as a secreted substrate. Similarly, deletion of the chaperone binding domains of IpgB1 and three additional Spa15-dependent effectors result in alleles that are no longer recognized as secreted substrates despite the presence of intact N-terminal secretion signal sequences. This is in contrast with MLD-containing effectors that bind class IA dedicated chaperones, as deletion of the MLD of these effectors alleviates the chaperone requirement for secretion. These observations indicate that at least for substrates of class IB chaperones, the chaperone-effector complex plays a major role in defining type 3 secreted proteins and highlight how a single region of an effector can play important roles both within prokaryotic and eukaryotic cells.

Highlights

Modulation of host signaling pathways by bacterial pathogens is critical as it enables these microorganisms to colonize and replicate within or in the vicinity of eukaryotic host cells
Fusion of the first 105 residues of IpgB1 to ELMO is sufficient to generate a protein that localizes to the plasma membrane and promotes the formation of membrane ruffles [5]. To test whether this region of IpgB1 encodes a membrane localization domain, we investigated whether fusion of the first 100 residues of IpgB1 to GFP is sufficient to redirect GFP to the yeast plasma membrane
We investigated whether the ten residues present within the chaperone binding domain of IpgB1 are necessary to direct its membrane localization

Summary

Introduction

Modulation of host signaling pathways by bacterial pathogens is critical as it enables these microorganisms to colonize and replicate within or in the vicinity of eukaryotic host cells. In this study, using the yeast Saccharomyces cerevisiae as a model system [12,13], we have identified a twenty amino acid region of IpgB1 responsible for its membrane localization This region, like that of the previously mapped membrane localization domains (MLDs) of several other type 3 effectors, is enriched in hydrophobic residues that likely promote membrane targeting. Included within these twenty residues are ten that define the IpgB1 chaperone binding domain (CBD) [11]. Our studies demonstrate how a single region of an effector, in this case IpgB1, can play important roles both in defining the protein as a bacterial secreted substrate as well as in promoting membrane localization once delivered into mammalian cells

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