Translocators YopB and YopD from Yersinia enterocolitica Form a Multimeric Integral Membrane Complex in Eukaryotic Cell Membranes

Montagner, Caroline; Arquint, Christian; Cornelis, Guy R.

doi:10.1128/jb.05555-11

Cited by 59 publications

(55 citation statements)

References 35 publications

(41 reference statements)

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“…1). Therefore, the molecular mass obtained using this model system is in good agreement with the molecular mass roughly estimated for Yersinia translocons isolated from erythrocyte membranes using blue native gel electrophoresis (600 Ϯ 100 kDa) (42). Taken together, single molecule fluorescence and ensemble experiments suggest a model where the interaction of PopD with PopB, presumably a dimer formation, leads to the assembly of unique hetero-oligomeric structures with defined stoichiometry on lipid bilayers.…”

Section: Discussionsupporting

confidence: 79%

Type 3 Secretion Translocators Spontaneously Assemble a Hexadecameric Transmembrane Complex

Romano¹,

Tang²,

Rossi³

et al. 2016

Journal of Biological Chemistry

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A type 3 secretion system is used by many bacterial pathogens to inject proteins into eukaryotic cells. Pathogens insert a translocon complex into the target eukaryotic membrane by secreting two proteins known as translocators. How these translocators form a translocon in the lipid bilayer and why both proteins are required remains elusive. Pseudomonas aeruginosa translocators PopB and PopD insert pores into membranes forming homo-or hetero-complexes of undetermined stoichiometry. Single-molecule fluorescence photobleaching experiments revealed that PopD formed mostly hexameric structures in membranes, whereas PopB displayed a bi-modal distribution with 6 and 12 subunits peaks. However, individually the proteins are not functional for effector translocation. We have found that when added together, the translocators formed distinct heterocomplexes containing 8 PopB and 8 PopD molecules. Thus, the interaction between PopB and PopD guide the assembly of a unique hetero-oligomer in membranes.The transport of proteins across membranes is essential at many stages of pathogen infection and colonization of human cells. This process typically involves the discharge of proteins from the pathogen (secretion) and the introduction of these secreted toxins/effectors into the cytosol of the target cell (translocation). Many pathogens, including the Shigella, Salmonella, Yersinia, and Pseudomonas species, exploit a sophisticated and efficient mechanism of protein secretion and translocation known as type III secretion (T3S) 3 system (1, 2). The T3S system is a syringe-like macromolecular machine formed by more than 20 different proteins organized in three major structures to span: (i) the inner bacterial membrane, the periplasmic space, and the outer bacterial membrane (the secreton); (ii) the extracellular space (the needle); and (iii) the host cellular membrane (the translocon) (3-6).A phylogenetic analysis of bacterial T3S systems based on conservation of their basal body ATPase indicates the presence of at least 7 families of T3S machines. The Pseudomonas aeruginosa genome encodes a single T3S system grouped within the Ysc family, named after the Yersinia spp. T3S system (the archetypical T3S system in this family) (7). The Ysc family includes pathogens like Yersinia pestis, Y. pseudotuberculosis, Y. enterocolitica, Bordetella pertussis, Vibrio parahemeolyticus, and P. aeruginosa, among others. The Ysc family shares structural similarity with the Inv-Mxi-Spa family of T3S systems, which includes the secretion systems used by Salmonella enterica and Shigella spp. (8).Great progress has been made in the structural characterization of the secreton and the needle for different T3S system families (9). However, little is known about how T3S-secreted proteins are translocated across the plasma membrane of the target cell to alter the normal function of the host (4). Two T3S-secreted proteins, known as the T3S translocators, insert into the target membrane to facilitate effector translocation. P. aeruginosa translocators PopB/PopD...

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Section: Discussionsupporting

confidence: 79%

Type 3 Secretion Translocators Spontaneously Assemble a Hexadecameric Transmembrane Complex

Romano¹,

Tang²,

Rossi³

et al. 2016

Journal of Biological Chemistry

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“…Translocon proteins form a hetero-oligomeric protein complex of presumably six to eight subunits with an internal diameter of approximately 1.2 to 3.5 nm (44,113,236,339,387,403,490,569). Recent crystal structure analysis of the translocon proteins IpaB from S. flexneri and SipB from Salmonella spp.…”

Section: Port Of Entry For Effector Proteins-the Translocon and The Tmentioning

confidence: 99%

Protein Export According to Schedule: Architecture, Assembly, and Regulation of Type III Secretion Systems from Plant- and Animal-Pathogenic Bacteria

Büttner

2012

Microbiol Mol Biol Rev

362

482

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SUMMARY Flagellar and translocation-associated type III secretion (T3S) systems are present in most Gram-negative plant- and animal-pathogenic bacteria and are often essential for bacterial motility or pathogenicity. The architectures of the complex membrane-spanning secretion apparatuses of both systems are similar, but they are associated with different extracellular appendages, including the flagellar hook and filament or the needle/pilus structures of translocation-associated T3S systems. The needle/pilus is connected to a bacterial translocon that is inserted into the host plasma membrane and mediates the transkingdom transport of bacterial effector proteins into eukaryotic cells. During the last 3 to 5 years, significant progress has been made in the characterization of membrane-associated core components and extracellular structures of T3S systems. Furthermore, transcriptional and posttranscriptional regulators that control T3S gene expression and substrate specificity have been described. Given the architecture of the T3S system, it is assumed that extracellular components of the secretion apparatus are secreted prior to effector proteins, suggesting that there is a hierarchy in T3S. The aim of this review is to summarize our current knowledge of T3S system components and associated control proteins from both plant- and animal-pathogenic bacteria.

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“…For translocation of Yops to occur, Yersinia must insert two proteins, YopB and YopD, into the membranes of targeted cells (1,22). YopD, but not YopB, has been found in purified needle preparations from Yersinia enterocolitica; however, its location on or within these needles has not be visualized by EM (10,11).…”

mentioning

confidence: 99%

“…While it has been speculated that the YopD in these complexes is due to secreted but insoluble YopD which copurifies with needles (11,12), recent work has also shown that YopD copurifies with LcrVcapped needles (10). YopB and YopD form a hetero-oligomeric pore in mammalian plasma membranes, termed the translocon, and are critical for the formation of a pore through which effector proteins are thought to travel (1,22). Purified YopB and YopD can insert into lipid bilayers (23,24), most likely because of their hydrophobic nature.…”

mentioning

confidence: 99%

A Mutant with Aberrant Extracellular LcrV-YscF Interactions Fails To Form Pores and Translocate Yop Effector Proteins but Retains the Ability To Trigger Yop Secretion in Response to Host Cell Contact

et al. 2013

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The plasmid-encoded type three secretion system (TTSS) of Yersinia spp. is responsible for the delivery of effector proteins into cells of the innate immune system, where these effectors disrupt the target cells' activity. Successful translocation of effectors into mammalian cells requires Yersinia to both insert a translocon into the host cell membrane and sense contact with host cells. To probe the events necessary for translocation, we investigated protein-protein interactions among TTSS components of the needle-translocon complex using a chemical cross-linking-based approach. We detected extracellular protein complexes containing YscF, LcrV, and YopD that were dependent upon needle formation. The formation of these complexes was evaluated in a secretion-competent but translocation-defective mutant, the YscFD28AD46A strain (expressing YscF with the mutations D28A and D46A). We found that one of the YscF and most of the LcrV and YopD cross-linked complexes were nearly absent in this mutant. Furthermore, the YscFD28AD46A strain did not support YopB insertion into mammalian membranes, supporting the idea that the LcrV tip complex is required for YopB insertion and translocon formation. However, the YscFD28AD46A strain did secrete Yops in the presence of host cells, indicating that a translocation-competent tip complex is not required to sense contact with host cells to trigger Yop secretion. In conclusion, in the absence of cross-linkable LcrV-YscF interactions, translocon insertion is abolished, but Yersinia still retains the ability to sense cell contact.

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Translocators YopB and YopD from Yersinia enterocolitica Form a Multimeric Integral Membrane Complex in Eukaryotic Cell Membranes

Cited by 59 publications

References 35 publications

Type 3 Secretion Translocators Spontaneously Assemble a Hexadecameric Transmembrane Complex

Type 3 Secretion Translocators Spontaneously Assemble a Hexadecameric Transmembrane Complex

Protein Export According to Schedule: Architecture, Assembly, and Regulation of Type III Secretion Systems from Plant- and Animal-Pathogenic Bacteria

A Mutant with Aberrant Extracellular LcrV-YscF Interactions Fails To Form Pores and Translocate Yop Effector Proteins but Retains the Ability To Trigger Yop Secretion in Response to Host Cell Contact

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