scholarly journals Holin-dependent secretion of the large clostridial toxin TpeL by Clostridium perfringens

2021 ◽  
Author(s):  
Angela Saadat ◽  
Stephen B. Melville
Keyword(s):  
Virulence Factors ◽  
Clostridium Perfringens ◽  
Signal Sequence ◽  
Secretion System ◽  
Membrane Topology ◽  
Secretion Signal ◽  
Clostridioides Difficile ◽  
Clostridium Novyi ◽  
Pass Through ◽  
Mutant Forms

Large clostridial toxins (LCTs) are secreted virulence factors found in several species, including Clostridioides difficile, Clostridium perfringens, Paeniclostridium sordellii, and Clostridium novyi. LCTs are large toxins that lack a secretion signal sequence and studies by others have shown the LCTs of C. difficile, TcdA and TcdB, require a holin-like protein, TcdE, for secretion. The TcdE gene is located on the PaLoc pathogenicity locus of C. difficile and holin-encoding genes are also present in the LCT-encoded PaLocs from P. sordellii and C. perfringens. However, the holin (TpeE) associated with the C. perfringens LCT, TpeL, has no homology and a different membrane topology than TcdE. In addition, TpeE has an identical membrane topology as the TatA protein, which is the core of the twin-arginine (Tat) secretion system. To determine if TpeE was necessary and sufficient to secrete TpeL, the genes from a type C strain of C. perfringens were expressed in a type A strain of C. perfringens, HN13, and secretion was measured using western blotting methods. We found that TpeE was required for TpeL secretion and secretion was not due to cell lysis. Mutant forms of TpeE lacking an amphipathic helix and a charged C-terminal domain failed to secrete TpeL and mutations that deleted conserved LCT domains in TpeL indicated only the full length protein could be secreted. In summary, we have identified a novel family of holin-like proteins that can function, in some cases, as a system of protein secretion for proteins that need to fold in the cytoplasm. Importance: Little is known about the mechanism by which LCTs are secreted. Since LCTs are major virulence factors in clostridial pathogens, we wanted to define the mechanism by which an LCT in C. perfringens, TpeL, is secreted by a protein (TpeE) lacking homology to previously described secretion-associated holins. We discovered TpeE is a member of widely dispersed class of holin proteins, and TpeE is necessary for secretion of TpeL. TpeE bears a high degree of similarity in membrane topology to TatA proteins, which form the pore through which Tat-secretion substrates pass through the cytoplasmic membrane. Thus, the TpeE-TpeL secretion system may be a model for understanding not only holin-dependent secretion, but also how TatA proteins function in the secretion process.

scholarly journals VirB4- and VirD4-like ATPases, components of a putative type 4C secretion system in Clostridioides difficile

2021 ◽  
Author(s):  
Julya Sorokina ◽  
Irina Sokolova ◽  
Ivan Rybolovlev ◽  
Natalya Shevlyagina ◽  
Vasiliy Troitskiy ◽  
...  
Keyword(s):  
Enzymatic Activity ◽  
Virulence Factors ◽  
Gene Clusters ◽  
Binding Activity ◽  
Secretion System ◽  
Dna Binding Activity ◽  
Dna And Rna ◽  
Clostridioides Difficile ◽  
The Stability ◽  
The One

The type 4 secretion system (T4SS) represents a bacterial nanomachine capable of trans-cell wall transportation of proteins and DNA and has attracted intense interest due to its roles in the pathogenesis of infectious diseases. In the current investigation, we uncovered three distinct gene clusters in Clostridioides difficile strain 630 encoding proteins structurally related to components of the VirB4/D4 type 4C secretion system from Streptococcus suis strain 05ZYH33 and located within sequences of conjugative transposons (CTn). Phylogenic analysis revealed that VirB4- and VirD4-like proteins of the CTn4 locus, on the one hand, and those of the CTn2 and CTn5 loci, on the other hand, fit into separate clades, suggesting specific roles of identified secretion system variants in the physiology of C. difficile . Our further study on VirB4- and VirD4-like products encoded by CTn4 revealed that both proteins possess Mg 2+ -dependent ATPase activity, form oligomers (most likely hexamers) in aqueous solutions, and rely on potassium but not sodium ions for the highest catalytic rate. VirD4 binds nonspecifically to DNA and RNA. The DNA-binding activity of VirD4 strongly decreased with the W241A variant. Mutations in the nucleotide sequences encoding presumable Walker A and Walker B motifs decreased the stability of the oligomers and significantly but not completely attenuated the enzymatic activity of VirB4. In VirD4, substitutions of amino acid residues in the peptides reminiscent of Walker structural motifs neither attenuated the enzymatic activity of the protein nor influenced the oligomerization state of the ATPase. Importance C. difficile is a gram-positive, anaerobic, spore-forming bacterium that causes life-threatening colitis in humans. Major virulence factors of the microorganism include the toxins TcdA, TcdB and CDT. However, other bacterial products, including a type 4C secretion system, have been hypothesized to contribute to the pathogenesis of the infection and are considered possible virulence factors of C. difficile . In the current paper, we describe the structural organization of putative T4SS machinery in C. difficile and characterize its VirB4- and VirD4-like components. Our studies, in addition to its significance for basic science, can potentially aid the development of antivirulence drugs suitable for the treatment of C. difficile infection.

Large Clostridial Toxins: Mechanisms and Roles in Disease

2021 ◽  
Author(s):  
Kathleen E. Orrell ◽  
Roman A. Melnyk
Keyword(s):  
Clostridium Perfringens ◽  
Target Cells ◽  
Content Type ◽  
Clostridioides Difficile ◽  
Clostridium Novyi

Large clostridial toxins (LCTs) are a family of bacterial exotoxins that infiltrate and destroy target cells. Members of the LCT family include Clostridioides difficile toxins TcdA and TcdB, Paeniclostridium sordellii toxins TcsL and TcsH, Clostridium novyi toxin TcnA, and Clostridium perfringens toxin TpeL.

scholarly journals Identification of new Dickeya dadantii virulence factors secreted by the type 2 secretion system

2021 ◽  
Author(s):  
Guy Condemine ◽  
Bastien Le Redout
Keyword(s):  
Pathogenic Bacteria ◽  
Signal Sequence ◽  
Variable Number ◽  
Secretion System ◽  
Dickeya Dadantii ◽  
Secretion Signal ◽  
Plant Infection ◽  
Wide Range ◽  
Type 2 Secretion

Dickeya are plant pathogenic bacteria able to provoke disease on a wide range of plants. A type 2 secretion system named Out is necessary for bacterial virulence. Its study in D. dadantii showed that it secretes a wide range of pectinolytic enzymes. However, the full repertoire of exoproteins it can secrete has not been identified. No secretion signal present on the protein allows the identification of substrates of a type 2 secretion system. To identify new Out substrates, we analyzed D. dadantii transcriptome data obtained in plant infection condition and searched for genes strongly induced encoding a protein with a signal sequence. We identified four new Out-secreted proteins: the expansin YoaJ, VirK and two proteins of the DUF 4879 family, SvfA and SvfB. We showed that SvfA and SvfB are required for full virulence of D. dadantii and showed that Svf proteins are present with a variable number of copies, up to three in D. fanghzongdai, in other Pectobacteriaceae. This work opens the way to the study of the role in virulence of non-pectinolytic proteins secreted by the Out pathway in Pectobacteriaceae.

scholarly journals Baicalin Represses Type Three Secretion System of Pseudomonas aeruginosa through PQS System

Molecules ◽  
2021 ◽  
Vol 26 (6) ◽  
pp. 1497
Author(s):  
Pansong Zhang ◽  
Qiao Guo ◽  
Zhihua Wei ◽  
Qin Yang ◽  
Zisheng Guo ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Virulence Factors ◽  
Mammalian Cells ◽  
Secretion System ◽  
Chinese Herbs ◽  
Infection Model ◽  
Lung Pathology ◽  
Promising Alternative ◽  
The Impact

Therapeutics that target the virulence of pathogens rather than their viability offer a promising alternative for treating infectious diseases and circumventing antibiotic resistance. In this study, we searched for anti-virulence compounds against Pseudomonas aeruginosa from Chinese herbs and investigated baicalin from Scutellariae radix as such an active anti-virulence compound. The effect of baicalin on a range of important virulence factors in P. aeruginosa was assessed using luxCDABE-based reporters and by phenotypical assays. The molecular mechanism of the virulence inhibition by baicalin was investigated using genetic approaches. The impact of baicalin on P. aeruginosa pathogenicity was evaluated by both in vitro assays and in vivo animal models. The results show that baicalin diminished a plenty of important virulence factors in P. aeruginosa, including the Type III secretion system (T3SS). Baicalin treatment reduced the cellular toxicity of P. aeruginosa on the mammalian cells and attenuated in vivo pathogenicity in a Drosophila melanogaster infection model. In a rat pulmonary infection model, baicalin significantly reduced the severity of lung pathology and accelerated lung bacterial clearance. The PqsR of the Pseudomonas quinolone signal (PQS) system was found to be required for baicalin’s impact on T3SS. These findings indicate that baicalin is a promising therapeutic candidate for treating P. aeruginosa infections.

scholarly journals Folding Control in the Path of Type 5 Secretion

Toxins ◽  
2021 ◽  
Vol 13 (5) ◽  
pp. 341
Author(s):  
Nathalie Dautin
Keyword(s):  
Protein Folding ◽  
Virulence Factors ◽  
Secretion System ◽  
Gram Negative Bacteria ◽  
Secretion Systems ◽  
Gram Negative ◽  
Final Destination ◽  
Cell Compartments ◽  
Recent Addition ◽  
Functionally Diverse

The type 5 secretion system (T5SS) is one of the more widespread secretion systems in Gram-negative bacteria. Proteins secreted by the T5SS are functionally diverse (toxins, adhesins, enzymes) and include numerous virulence factors. Mechanistically, the T5SS has long been considered the simplest of secretion systems, due to the paucity of proteins required for its functioning. Still, despite more than two decades of study, the exact process by which T5SS substrates attain their final destination and correct conformation is not totally deciphered. Moreover, the recent addition of new sub-families to the T5SS raises additional questions about this secretion mechanism. Central to the understanding of type 5 secretion is the question of protein folding, which needs to be carefully controlled in each of the bacterial cell compartments these proteins cross. Here, the biogenesis of proteins secreted by the Type 5 secretion system is discussed, with a focus on the various factors preventing or promoting protein folding during biogenesis.

Saccharomyces cerevisiae secretes and correctly processes human interferon hybrid proteins containing yeast invertase signal peptides

1986 ◽  
Vol 6 (5) ◽  
pp. 1812-1819
Author(s):  
C N Chang ◽  
M Matteucci ◽  
L J Perry ◽  
J J Wulf ◽  
C Y Chen ◽  
...  
Keyword(s):  
Amino Acid ◽  
Signal Sequence ◽  
Phosphoglycerate Kinase ◽  
Yeast Cells ◽  
Signal Peptidase ◽  
Human Interferon ◽  
Expression Plasmid ◽  
Kinase Gene ◽  
Secretion Signal ◽  
Yeast Invertase

Synthetic oligonucleotides coding for the yeast invertase secretion signal peptide were fused to the gene for the mature form of human interferon (huIFN-alpha 2). Two plasmids (E3 and F2) were constructed. E3 contained the invertase signal codons in a reading frame with the mature huIFN-alpha 2 gene. F2 had a deletion of the codon for alanine at amino acid residue-5 in the invertase signal and an addition of a methionine codon located between the coding sequences for the invertase signal and mature huIFN-alpha 2. Both hybrid genes were located adjacent to the promoter from the 3-phosphoglycerate kinase gene on the multicopy yeast expression plasmid, YEp1PT. Yeast transformants containing these plasmids produced somewhat more IFN than did the same expression plasmid containing the IFN gene with its human secretion signal sequence. HuIFN-alpha 2, purified from the medium of yeast cells containing E3, was found to be processed at the correct site. The huIFN-alpha 2 made by plasmid F2 was found to be completely processed at the junction between the invertase signal (a variant) and the methionine of methionine-huIFN-alpha 2. These results strongly suggested that the invertase signal (or its variant) attached to huIFN was efficiently recognized by the presumed signal recognition particle and was cleaved by the signal peptidase in the yeast cells. These results also suggested that amino acid changes on the right side of the cleavage site did not necessarily prevent cleavage or secretion.

scholarly journals Cloning of fibA, Encoding an Immunogenic Subunit of the Fibril-Like Surface Structure ofPeptostreptococcus micros

1999 ◽  
Vol 181 (8) ◽  
pp. 2485-2491 ◽  
Author(s):  
B. H. A. Kremer ◽  
J. J. E. Bijlsma ◽  
J. G. Kusters ◽  
J. de Graaff ◽  
T. J. M. van Steenbergen
Keyword(s):  
Amino Acid ◽  
Surface Structure ◽  
Signal Sequence ◽  
Open Reading Frames ◽  
Periodontal Pathogen ◽  
Immunogold Electron Microscopy ◽  
Expression Library ◽  
Gram Positive ◽  
Secretion Signal ◽  
Specific Serum

ABSTRACT Although we are currently unaware of its biological function, the fibril-like surface structure is a prominent characteristic of the rough (Rg) genotype of the gram-positive periodontal pathogenPeptostreptococcus micros. The smooth (Sm) type of this species as well as the smooth variant of the Rg type (RgSm) lack these structures on their surface. A fibril-specific serum, as determined by immunogold electron microscopy, was obtained through adsorption of a rabbit anti-Rg type serum with excess bacteria of the RgSm type. This serum recognized a 42-kDa protein, which was subjected to N-terminal sequencing. Both clones of a λTriplEx expression library that were selected by immunoscreening with the fibril-specific serum contained an open reading frame, designatedfibA, encoding a 393-amino-acid protein (FibA). The 15-residue N-terminal amino acid sequence of the 42-kDa antigen was present at positions 39 to 53 in FibA; from this we conclude that the mature FibA protein contains 355 amino acids, resulting in a predicted molecular mass of 41,368 Da. The putative 38-residue signal sequence of FibA strongly resembles other gram-positive secretion signal sequences. The C termini of FibA and two open reading frames directly upstream and downstream of fibA exhibited significant sequence homology to the C termini of a group of secreted and surface-located proteins of other gram-positive cocci that are all presumably involved in anchoring of the protein to carbohydrate structures. We conclude that FibA is a secreted and surface-located protein and as such is part of the fibril-like structures.

scholarly journals Novel Drivers of Virulence in Clostridioides difficile Identified via Context-Specific Metabolic Network Analysis

mSystems ◽  
2021 ◽  
Author(s):  
Matthew L. Jenior ◽  
Jhansi L. Leslie ◽  
Deborah A. Powers ◽  
Elizabeth M. Garrett ◽  
Kimberly A. Walker ◽  
...  
Keyword(s):  
Network Analysis ◽  
Metabolic Network ◽  
Virulence Factors ◽  
Metabolic Pathways ◽  
Content Type ◽  
Hospital Acquired Infections ◽  
Metabolic Network Analysis ◽  
Clostridioides Difficile ◽  
Context Specific ◽  
Hospital Acquired

Clostridioides difficile has become the leading single cause of hospital-acquired infections. Numerous studies have demonstrated the importance of specific metabolic pathways in aspects of C. difficile pathophysiology, from initial colonization to regulation of virulence factors.

scholarly journals Non-O1/Non-O139 Vibrio cholerae Carrying Multiple Virulence Factors and V. cholerae O1 in the Chesapeake Bay, Maryland

2015 ◽  
Vol 81 (6) ◽  
pp. 1909-1918 ◽  
Author(s):  
Daniela Ceccarelli ◽  
Arlene Chen ◽  
Nur A. Hasan ◽  
Shah M. Rashed ◽  
Anwar Huq ◽  
...  
Keyword(s):  
Chesapeake Bay ◽  
Vibrio Cholerae ◽  
Virulence Factors ◽  
Fluorescent Antibody ◽  
Secretion System ◽  
Surveillance Program ◽  
Toxin Gene ◽  
Protease Gene ◽  
Content Type ◽  
Pathogenic Variants

ABSTRACTNon-O1/non-O139Vibrio choleraeinhabits estuarine and coastal waters globally, but its clinical significance has not been sufficiently investigated, despite the fact that it has been associated with septicemia and gastroenteritis. The emergence of virulent non-O1/non-O139V. choleraeis consistent with the recognition of new pathogenic variants worldwide. Oyster, sediment, and water samples were collected during a vibrio surveillance program carried out from 2009 to 2012 in the Chesapeake Bay, Maryland.V. choleraeO1 was detected by a direct fluorescent-antibody (DFA) assay but was not successfully cultured, whereas 395 isolates of non-O1/non-O139V. choleraewere confirmed by multiplex PCR and serology. Only a few of the non-O1/non-O139V. choleraeisolates were resistant to ampicillin and/or penicillin. Most of the isolates were sensitive to all antibiotics tested, and 77 to 90% carried the El Tor variant hemolysin genehlyAET, the actin cross-linking repeats in toxin genertxA, the hemagglutinin protease genehap, and the type 6 secretion system. About 19 to 21% of the isolates carried the neuraminidase-encoding genenanHand/or the heat-stable toxin (NAG-ST), and only 5% contained a type 3 secretion system. None of the non-O1/non-O139V. choleraeisolates containedVibriopathogenicity island-associated genes. However,ctxA,ace, orzotwas present in nine isolates. Fifty-five different genotypes showed up to 12 virulence factors, independent of the source of isolation, and represent the first report of both antibiotic susceptibility and virulence associated with non-O1/non-O139V. choleraefrom the Chesapeake Bay. Since these results confirm the presence of potentially pathogenic non-O1/non-O139V. cholerae, monitoring for totalV. cholerae, regardless of serotype, should be done within the context of public health.

Fecal PCR testing for detection of Clostridium perfringens and Clostridioides difficile toxin genes and other pathogens in foals with diarrhea: 28 cases

2021 ◽  
pp. 104063872110475
Author(s):  
K. Gary Magdesian ◽  
Samantha Barnum ◽  
Nicola Pusterla
Keyword(s):  
Blood Cell ◽  
Clostridium Perfringens ◽  
Red Blood Cell ◽  
Total Protein ◽  
Toxin Gene ◽  
Significant Morbidity ◽  
Gene Sequences ◽  
Clostridioides Difficile ◽  
Gastrointestinal Pathogens ◽  
Cell Concentrations

Clostridium perfringens and Clostridioides difficile cause significant morbidity and mortality in foals. Antemortem diagnosis of C. perfringens infection has been complicated by a paucity of tests available for toxin detection. Fecal PCR panels have assays for a variety of C. perfringens toxin gene sequences as well as for several other foal gastrointestinal pathogens. We evaluated results of a comprehensive fecal diarrhea PCR panel in 28 foals that had been presented to a referral hospital because of diarrhea. Sixteen (57%) foals were positive for C. perfringens and/or C. difficile toxin gene sequences on fecal PCR, including 3 foals positive for NetF toxin. These foals were younger ( p = 0.0029) and had higher hematocrits ( p = 0.0087), hemoglobin ( p = 0.0067), and red blood cell concentrations ( p = 0.028) than foals with diarrhea that tested negative for clostridial toxins. The foals had lower total protein concentrations ( p = 0.045) and were more likely to have band neutrophils on a CBC ( p = 0.013; OR: 16.2). All 3 foals with NetF toxin gene sequences detected in feces survived to discharge, indicating that diarrhea caused by NetF toxigenic C. perfringens isolates is not uniformly fatal.

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