Exoproteomic analysis of the SecA2-dependent secretion in Listeria monocytogenes EGD-e

Rénier, Sandra; Chambon, Christophe; Viala, Didier; Chagnot, Caroline; Hébraud, Michel; Desvaux, Mickaël

doi:10.1016/j.jprot.2012.11.027

Cited by 38 publications

(38 citation statements)

References 68 publications

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“…In order not to compete for substrate, SecA2 is bound to ADP in a dormant state until it is required (71). Considering there is no conservation in the signal peptides of proteins being secreted through the SecA2-dependent pathway in L. monocytogenes, it remains unclear how proteins are targeted to SecA2 (14,32) and how that can be overcome in certain conditions.…”

Section: Discussionmentioning

confidence: 99%

“…Mutants in secA2 exhibit a chaining phenotype and are approximately 1,000-fold less virulent in animal models of infection (14,25,28). These mutants secrete a reduced subset of proteins, including two major autolysins, P60 (CwhA) and NamA (MurA), that contribute to septation and pathogenesis (14,(29)(30)(31)(32). To better understand SecA2-dependent protein secretion and the contribution that SecA2 makes to L. monocytogenes pathogenesis, we undertook a suppressor analysis of a ⌬secA2 mutant.…”

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confidence: 99%

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A prl Mutation in SecY Suppresses Secretion and Virulence Defects of Listeria monocytogenes secA2 Mutants

2015

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The bulk of bacterial protein secretion occurs through the conserved SecY translocation channel that is powered by SecA-dependent ATP hydrolysis. Many Gram-positive bacteria, including the human pathogen Listeria monocytogenes, possess an additional nonessential specialized ATPase, SecA2. SecA2-dependent secretion is required for normal cell morphology and virulence in L. monocytogenes; however, the mechanism of export via this pathway is poorly understood. L. monocytogenes secA2 mutants form rough colonies, have septation defects, are impaired for swarming motility, and form small plaques in tissue culture cells. In this study, 70 spontaneous mutants were isolated that restored swarming motility to L. monocytogenes secA2 mutants. Most of the mutants had smooth colony morphology and septated normally, but all were lysozyme sensitive. Five representative mutants were subjected to whole-genome sequencing. Four of the five had mutations in proteins encoded by the lmo2769 operon that conferred lysozyme sensitivity and increased swarming but did not rescue virulence defects. A point mutation in secY was identified that conferred smooth colony morphology to secA2 mutants, restored wild-type plaque formation, and increased virulence in mice. This secY mutation resembled a prl suppressor known to expand the repertoire of proteins secreted through the SecY translocation complex. Accordingly, the ⌬secA2prlA1 mutant showed wild-type secretion levels of P60, an established SecA2-dependent secreted autolysin. Although the prl mutation largely suppressed almost all of the measurable SecA2-dependent traits, the ⌬secA2prlA1 mutant was still less virulent in vivo than the wild-type strain, suggesting that SecA2 function was still required for pathogenesis.T he essential general secretory (Sec) pathway is responsible for exporting the majority of secreted proteins across the bacterial cell membrane (1-3). Much of what we know about this pathway was discovered in Escherichia coli by using genetic screens to identify loss-of-function mutations in components of the Sec system. This led to the identification of components of the SecYEG complex that forms the translocation channel and the SecA ATPase that binds to signal sequences to drive the export of precursor proteins across the channel (3-9). In contrast to loss-of-function sec mutations, prl alleles are gain-of-function mutations, which expand the repertoire of substrates being exported across the SecYEG channel (6-8, 10, 11). The most dominant prl variants are in secY, known as prlA mutations, which allow for the secretion of proteins with altered signal sequence (7, 9, 11) without significantly altering the secretion of other proteins (6,12).In addition to the canonical SecA, an accessory cytosolic ATPase, SecA2, has been identified in Mycobacterium and a subset of other Gram-positive bacteria. Unlike SecA, SecA2 is not essential for cell viability but is required for virulence in some pathogenic bacteria (13-15). SecA1 and SecA2 are homologous but are not interchangeable (16)...

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Section: Discussionmentioning

confidence: 99%

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confidence: 99%

A prl Mutation in SecY Suppresses Secretion and Virulence Defects of Listeria monocytogenes secA2 Mutants

2015

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“…SecA2 was shown to facilitate the secretion of several autolytic enzymes, cell wall proteins, and a superoxide dismutase, termed MnSOD, all of which contribute to L. monocytogenes in vivo infection (13,24,25). PrsA2 was shown to promote the folding and activity of LLO and PC-PLC upon secretion (26)(27)(28)(29); however, the role of SecDF in L. monocytogenes has not been characterized.…”

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confidence: 99%

Membrane Chaperone SecDF Plays a Role in the Secretion of Listeria monocytogenes Major Virulence Factors

Tamar

Pozniak

Rabinovich

et al. 2013

Journal of Bacteriology

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Listeria monocytogenes is a Gram-positive human intracellular pathogen that infects diverse mammalian cells. Upon invasion, L. monocytogenes secretes multiple virulence factors that target host cellular processes and promote infection. It has been presumed, but was not empirically established, that the Sec translocation system is the primary mediator of this secretion. Here, we validate an important role for SecDF, a component of the Sec system, in the secretion of several critical L. monocytogenes virulence factors. A ⌬secDF mutant is demonstrated to exhibit impaired membrane translocation of listeriolysin O (LLO), PlcA, PlcB, and ActA, factors that mediate L. monocytogenes phagosomal escape and spread from cell to cell. This impaired translocation was monitored by accumulation of the factors on the bacterial membrane and by reduced activity upon secretion. This defect in secretion is shown to be associated with a severe intracellular growth defect of the ⌬secDF mutant in macrophages and a less virulent phenotype in mice, despite normal growth in laboratory medium. We further show that SecDF is upregulated when the bacteria reside in macrophage phagosomes and that it is necessary for efficient phagosomal escape. Taken together, these data support the premise that SecDF plays a role as a chaperone that facilitates the translocation of L. monocytogenes virulence factors during infection.

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“…Mycobacterial SecA2 recognizes the mature domains of a few secretory precursors to assist in their secretion, suggesting its primary function may be maintenance of export competence for proteins with the propensity for rapid folding (46). Listeria monocytogenes secA2 was identified because mutations in the structural gene affect colony shape and cellular morphology, a phenotype that is attributed to the diminished secretion of cell wall hydrolases (i.e., p60 autolysin, NamA hydrolase, and two dozen other substrates) (47)(48)(49). The gene for the Listeria SecA2 p60 substrate, cwhA, is located immediately adjacent to secA2, and this locus is conserved among pathogenic and nonpathogenic Listeria species (50).…”

Section: Discussionmentioning

confidence: 99%

Bacillus anthracis SlaQ Promotes S-Layer Protein Assembly

Nguyen-Mau

Schneewind

et al. 2015

J Bacteriol

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Bacillus anthracis vegetative forms assemble an S-layer comprised of two S-layer proteins, Sap and EA1. A hallmark of S-layer proteins are their C-terminal crystallization domains, which assemble into a crystalline lattice once these polypeptides are deposited on the bacterial surface via association between their N-terminal S-layer homology domains and the secondary cell wall polysaccharide. Here we show that slaQ, encoding a small cytoplasmic protein conserved among pathogenic bacilli elaborating S-layers, is required for the efficient secretion and assembly of Sap and EA1. S-layer protein precursors cosediment with SlaQ, and SlaQ appears to facilitate Sap assembly. Purified SlaQ polymerizes and when mixed with purified Sap promotes the in vitro formation of tubular S-layer structures. A model is discussed whereby SlaQ, in conjunction with S-layer secretion factors SecA2 and SlaP, promotes localized secretion and S-layer assembly in B. anthracis. IMPORTANCES-layer proteins are endowed with the propensity for self-assembly into crystalline arrays. Factors promoting S-layer protein assembly have heretofore not been reported. We identified Bacillus anthracis SlaQ, a small cytoplasmic protein that facilitates S-layer protein assembly in vivo and in vitro.

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Exoproteomic analysis of the SecA2-dependent secretion in Listeria monocytogenes EGD-e

Cited by 38 publications

References 68 publications

A prl Mutation in SecY Suppresses Secretion and Virulence Defects of Listeria monocytogenes secA2 Mutants

A prl Mutation in SecY Suppresses Secretion and Virulence Defects of Listeria monocytogenes secA2 Mutants

Membrane Chaperone SecDF Plays a Role in the Secretion of Listeria monocytogenes Major Virulence Factors

Bacillus anthracis SlaQ Promotes S-Layer Protein Assembly

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