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StcE binds erythrocyte surfaces. (A) 250 ng StcE-His or Alexa-StcE-His was incubated with opsonized sheep erythrocytes for 10 min at 37C, after which the cells were washed and the binding of StcE was detected in a flow cytometer. (B) Increasing concentrations of AlexaStcE-His (0.1-25.6 g) were added to sheep erythrocytes. StcE binding was detected by flow cytometry as described above, and the geometric mean fluorescence for each sample was plotted against the corresponding concentration of Alexa-StcEHis.
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The complement system is an essential component of host defense against pathogens. Previous research in our laboratory identified StcE, a metalloprotease secreted by Escherichia coli O157:H7 that cleaves the serpin C1 esterase inhibitor (C1-INH), a major regulator of the classical complement cascade. Analyses of StcE-treated C1-INH activity reveale...
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... primary amines with the Alexa Fluor 488 dye (Alexa-StcE-His) were added to opsonized sheep erythrocytes for 10 min at 37C. Erythrocytes were pelleted and washed with VBS 2 before analysis by flow cytometry. Erythrocytes treated with Alexa-labeled StcE-His showed 80-fold greater mean fluorescence compared with cells treated with unlabeled StcE-His (Fig. 3 A), demonstrating a direct interaction be- tween these cells and the protease. Furthermore, we ob- served that StcE continues to bind sheep erythrocytes even at lower temperatures (0-4C), suggesting that this interac- tion is not mediated by an active cellular process (not de- picted). To determine if the interaction between erythro- ...
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... process (not de- picted). To determine if the interaction between erythro- cytes and StcE is specific and therefore saturable, we mixed increasing concentrations of Alexa-labeled StcE-His with 10 7 sheep erythrocytes as described above. We observed that this number of erythrocytes becomes saturated with StcE-His at 3.2 g of the protease in 500 l (Fig. 3 B). Based on the calculated molecular weight of StcE-His, at this concentration we estimate 1.8 10 6 molecules of StcE are bound per ...
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... For instance, desialylated C1-INH is thought to interact with hepatic asialoglycoprotein receptors (35), whereas C1-INH-C1s complexes bind to the low density lipoprotein receptor-related protein, LRP (36). Rather than modifying C1-INH in this way, however, our data indicate that StcE sequesters C1-INH to erythrocytes by itself binding to cells (Fig. 3) and acting as a "bridge" be- tween the serpin and the cell surface (Fig. 5). Indeed, the cleavage of C1-INH by StcE is unnecessary to provide in- creased protection against complement activity, as a point mutant of StcE defective in proteolytic activity against C1- INH protects erythrocytes to similar levels as wild-type StcE in vitro ...
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... to provide in- creased protection against complement activity, as a point mutant of StcE defective in proteolytic activity against C1- INH protects erythrocytes to similar levels as wild-type StcE in vitro (Fig. 5 A). The interaction between StcE and eryth- rocytes is specific, saturating the cells at 1.8 10 6 mole- cules of StcE per cell (Fig. 3 B). In turn, this allows a high affinity interaction between C1-INH and erythrocytes, reaching 2.25 10 6 molecules of C1-INH per cell in the presence of 1 g StcE. This amount of C1-INH (0.1 IU) is well within the physiological concentration of C1-INH found in serum, suggesting that this interaction might be bi- ologically relevant in ...
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Citations
... A common strategy used by bacteria to e v ade complement is the recruitment of complement inhibitors that regulate autologous complement activation to prevent inflammation and pathologies. For example, E. coli secretes the metalloprotease StcE that recruits and potentiates C1-INH, a serine protease inhibitor that negativ el y r egulates C1r, C1s (CP), MASP-1, and MASP-2 (LP) on host cells, pr otecting adher ent bacteria fr om complement (Lathem et al. 2004 ). ...
Bloodstream infection is a major public health concern associated with high mortality and high healthcare costs worldwide. Bacteremia can trigger fatal sepsis whose prevention, diagnosis and management have been recognized as a global health priority by the World Health Organization. Additionally, infection control is increasingly threatened by antimicrobial resistance, which is the focus of global action plans in the framework of a One Health response. In-depth knowledge of the infection process is needed to develop efficient preventive and therapeutic measures. The pathogenesis of bloodstream infection is a dynamic process resulting from the invasion of the vascular system by bacteria, which finely regulate their metabolic pathways and virulence factors to overcome the blood immune defenses and proliferate. In this review, we highlight our current understanding of determinants of bacterial survival and proliferation in the bloodstream and discuss their interactions with the molecular and cellular components of blood.
... Another important putative virulence factor is zinc metalloprotease (StcE) produced by the O157:H7 strain of EHEC 92 , released by the Type-II secretion system that is coded on pO157 virulence plasmid, which is offering great support for mammalian colonization by proteolytically exposing the cell surface of the host leading to close adherence of the organism to the host cell. 74,93 The present representation for the action of StcE states that primarily, StcE permits the movement of the bacterium along the oral cavity by splitting the mucin-type glycoprotein present in saliva utilizing its metalloprotease-mediated mucinase activity that is accountable for the aggregation of bacteria. By splitting glycoprotein 340 as well as mucin existing in saliva, it can break down and lessen the thickness of the mucus layer. ...
The Shigatoxigenic Escherichia coli (STEC) are bacterial enteropathogens responsible for some intensive clinical syndromes such as bloody diarrhoea, hemorrhagic colitis, hemolytic uremic syndrome, thrombotic thrombocytopenic purpura, and renal failure. These pathotypes come under the Enterohemorrhagic Escherichia coli (EHEC) group. Monogastric farm animals such as pigs, horses, chickens, ducks, turkeys and aquatic animals like shellfish, fishes, and wild animals can act as major spillover hosts of STEC strains and could serve as the potential source of infection. The pathogen is notorious as a quickly emergent strain with acquired characteristics like different variants of Shigatoxin, many antibiotic degrading enzymes, Intimin, Enterohemolysin, Auto-agglutination Adhesins, Catalase-peroxidase, Zinc metalloprotease, Subtilase cytotoxin, tolerance to multiple adverse conditions, and biofilm formation. The bacteria are known for its long survival in different adverse physical-chemical conditions. The formation of biofilm is one of the major factors responsible for their persistence. Multidrug resistance is another related trait contributing to the high mortality rate of these strains. STEC strains are good candidates for studying the emergence of pathogens with acquired characteristics like genes. In this article, various virulent traits and multidrug resistance that enabled the strain to emerge as a serious public health menace were reviewed.
... Another mechanism adopted by some pathogens to interfere with the complement system's initial steps is the acquisition of regulators involved in this stage, such as C1-INH. For instance, E. coli O157:H7 produces the secreted protease of C1 esterase inhibitor (StcE), which cleaves C1-INH and potentiates its inhibitory effect, thus downregulating the activation of both the CP and LP (44). B. recurrentis, the causative agent of louse-borne relapsing fever, is able to acquire the complement regulator C1-INH by the surface protein complement inhibition (CihC) (41). ...
The complement system is a fundamental part of the innate immune system that plays a key role in the battle of the human body against invading pathogens. Through its three pathways, represented by the classical, alternative, and lectin pathways, the complement system forms a tightly regulated network of soluble proteins, membrane-expressed receptors, and regulators with versatile protective and killing mechanisms. However, ingenious pathogens have developed strategies over the years to protect themselves from this complex part of the immune system. This review briefly discusses the sequence of the complement activation pathways. Then, we present a comprehensive updated overview of how the major four pathogenic groups, namely, bacteria, viruses, fungi, and parasites, control, modulate, and block the complement attacks at different steps of the complement cascade. We shed more light on the ability of those pathogens to deploy more than one mechanism to tackle the complement system in their path to establish infection within the human host.
... As a basis of capsular polysaccharides (CPS) antigenicity, S. suis is classified in 29 serotypes (1/2, 1-19, 21, 23-25 and 27-31) [3]. The serotype 1,2,5,9,14,16,28, and 31 can infect humans, and serotype 2 (SS2) is the most virulent and prevalent one in swine and humans worldwide [4]. Since the first reported cases of human S. suis infection in Denmark in 1968, more than 1600 cases have been reported worldwide [5]. ...
... Microbes have developed several mechanisms to disrupt the stringent cascade network of complement system to survive in the host. The bacterial complement evasion strategies are divided into five groups: (a) mimicking of complement regulatory proteins like the CD-59 like protein in B. burgdorferi [12]; (b) secretion of proteases to cleave complement components like C3degrading protein SpeB in S. pyogenes [13]; (c) complement exploitation for subverting the host response like the inhibition of TLR2 activity by inducing CR3 signaling on macrophages by F. tularensis [14]; (d) secretion of small bacterial proteins with complement inhibitory properties [15]; (e) recruiting of fluid phase complement regulatory proteins C1-INH [16], C4BP [17] and fH [18]. Over the past 20 years, a number of virulence factors are found to contribute to S. suis invasion of the epithelial barrier, immune evasion and persistence in the bloodstream and multiple host organs [3,19]. ...
Streptococcus suis (S. suis), a significant zoonotic bacterial pathogen impacting swine and human, is associated with severe systemic diseases such as streptococcal toxic shock-like syndrome, meningitis, septicaemia, and abrupt fatality. The multifaceted roles of complement components C5a and C3a extend to orchestrating inflammatory cells recruitment, oxidative burst induction, and cytokines release. Despite the pivotal role of subtilisin-like serine proteases in S. suis pathogenicity, their involvement in immune evasion remains underexplored. In the present study, we identify two cell wall-anchored subtilisin-like serine proteases in S. suis, SspA-1 and SspA-2, as binding partners for C3a and C5a. Through Co-Immunoprecipitation, Enzyme-Linked Immunosorbent and Far-Western Blotting Assays, we validate their interactions with the aforementioned components. However, SspA-1 and SspA-2 have no cleavage activity against complement C3a and C5a performed by Cleavage assay. Chemotaxis assays reveal that recombinant SspA-1 and SspA-2 effectively attenuate monocyte chemotaxis towards C3a and C5a. Notably, the ΔsspA-1, ΔsspA-1, and ΔsspA-1/2 mutant strains exhibit compromised survival in blood, and resistance of opsonophagocytosis, alongside impaired survival in blood and in vivo colonization compared to the parental strain SC-19. Critical insights from the murine and Galleria mellonella larva infection models further underscore the significance of sspA-1 in altering mortality rates. Collectively, our findings indicate that SspA-1 and SspA-2 are novel binding proteins for C3a and C5a, thereby shedding light on their pivotal roles in S. suis immune evasion and the pathogenesis.
... Interestingly, some pathogens purposefully activate the complement system in order to assist in the causation of disease, which is sometimes known as the "Hitchhiking principle" [46]. This principle is used by intracellular pathogens such as Mycobacterium tuberculosis and many viruses whereby they recruit a complement protein to their membrane or surface in order to enter host cells [47,48]. ...
... Lathem et al. [46] discovered that the secreted StcE metalloprotease of Escherichia coli is capable of binding C1INH. It was also found that StcE is capable of cleaving C1INH, which likely relates to its ability to inhibit the contact activation pathway, which is involved in coagulation and leads to the formation of thrombin or blood clots that prevent any further spreading of bacterial infections [49]. ...
Resistance to antibiotics in Bacteria is one of the biggest threats to human health. After decades of attempting to isolate or design antibiotics with novel mechanisms of action against bacterial pathogens, few approaches have been successful. Antibacterial drug discovery is now moving towards targeting bacterial virulence factors, especially immune evasion factors. Gram-negative bacteria present some of the most significant challenges in terms of antibiotic resistance. However, they are also able to be eliminated by the component of the innate immune system known as the complement system. In response, Gram-negative bacteria have evolved a variety of mechanisms by which they are able to evade complement and cause infection. Complement resistance mechanisms present some of the best novel therapeutic targets for defending against highly antibiotic-resistant pathogenic bacterial infections.
... In addition to well-characterized LEE-encoded and non-LEE-encoded regulators that control AE phenotype, a total of 10 fimbrial genes and 13 non-fimbrial adhesins were found in EHEC (Hayashi et al., 2001;Perna et al., 2001). Of which, the long polar fimbriae ( Jordan et al., 2004;Farfan et al., 2011;Lloyd et al., 2012), E. coli common plus (Rendon et al., 2007), the autotransporter protein, EhaG (Totsika et al., 2012), and the pO157 virulence plasmid-encoded zinc metalloprotease (StcE) (Lathem et al., 2004;Grys et al., 2005) are involved in EHEC adhesion to epithelial cells and host colonization Figures 1 and 2). ...
Enterohemorrhagic Escherichia coli (EHEC) is one of the most common E. coli pathotypes reported to cause several outbreaks of foodborne illnesses. EHEC is a zoonotic pathogen, and ruminants, especially cattle, are considered important reservoirs for the most common EHEC serotype, E. coli O157:H7. Humans are infected indirectly through the consumption of food (milk, meat, leafy vegetables, and fruits) and water contaminated by animal feces or direct contact with carrier animals or humans. E. coli O157:H7 is one of the most frequently reported causes of foodborne illnesses in developed countries. It employs two essential virulence mechanisms to trigger damage to the host. These are the development of attaching and effacing (AE) phenotypes on the intestinal mucosa of the host and the production of Shiga toxin (Stx) that causes hemorrhagic colitis and hemolytic uremic syndrome. The AE phenotype is controlled by the pathogenicity island, the locus of enterocyte effacement (LEE). The induction of both AE and Stx is under strict and highly complex regulatory mechanisms. Thus, a good understanding of these mechanisms, major proteins expressed, and environmental cues involved in the regulation of the expression of the virulence genes is vital to finding a method to control the colonization of reservoir hosts, especially cattle, and disease development in humans. This review is a concise account of the current state of knowledge of virulence gene regulation in the LEE-positive EHEC.
... The regulators of complement pathways C1-inhibitor (C1-INH) (Davis et al. 1986), C4b-binding protein (C4BP), MBL/ficolin/CLassociated protein-1 (MAP-1), and small MBL-associated protein (sMAP) (Schmidt et al. 2016) are utilized by various pathogens for their survival. Escherichia coli and Bordetella pertussis use C1-INH (Lathem et al. 2004;Marr et al. 2007), many pathogens exploit C4BP as part of their survival strategy Ram 2008, Hovingh et al. 2016). As coagulation inhibitors can affect LP, such as anti-thrombin inactivation of MASPs (Presanis et al. 2004), it may also be part of the survival strategy of various pathogens. ...
From time immemorial, natural products (NPs) are a major source of drugs in the pharmaceutical industry ranging from antibiotics, anti-depressants, anti-inflammatory, anti-tumor, and anti-cancer agents to name a few. Actinobacteria and their biosynthetic pathways contribute substantially to these drugs present in the laboratory level, clinical phase trials, and in the market. The majority of the secondary metabolites secreted are produced from the nonribosomal peptide synthetase (NRPS) and polyketide synthetase (PKS) enzymes. Apart from the nonribosomal peptides produced, ribosomally produced peptides that undergo drastic post-translational modifications also play a part in contributing to the essential drugs produced. The chapter deals with both nonribosomal peptides (NRPs) and ribosomally synthesized and post-translationally modified peptides (RiPPs).
... C1-inh can also be appropriated by pathogens to prevent initiation of the classical and lectin pathways. The first report of C1-inh recruitment by a pathogen appeared in 2004, when the ability of the E. coli protein StcE to bind C1-inh to host cell surfaces was described (Lathem et al., 2004). The importance of this immune evasion strategy is highlighted by the fact that virulent Bordetella pertussis binds C1-inh whereas avirulent Bordetella spp. ...
The genus Burkholderia contains over 80 different Gram-negative species including both plant and human pathogens, the latter of which can be classified into one of two groups: the Burkholderia pseudomallei complex (Bpc) or the Burkholderia cepacia complex (Bcc). Bpc pathogens Burkholderia pseudomallei and Burkholderia mallei are highly virulent, and both have considerable potential for use as Tier 1 bioterrorism agents; thus there is great interest in the development of novel vaccines and therapeutics for the prevention and treatment of these infections. While Bcc pathogens Burkholderia cenocepacia, Burkholderia multivorans, and Burkholderia cepacia are not considered bioterror threats, the incredible impact these infections have on the cystic fibrosis community inspires a similar demand for vaccines and therapeutics for the prevention and treatment of these infections as well. Understanding how these pathogens interact with and evade the host immune system will help uncover novel therapeutic targets within these organisms. Given the important role of the complement system in the clearance of bacterial pathogens, this arm of the immune response must be efficiently evaded for successful infection to occur. In this review, we will introduce the Burkholderia species to be discussed, followed by a summary of the complement system and known mechanisms by which pathogens interact with this critical system to evade clearance within the host. We will conclude with a review of literature relating to the interactions between the herein discussed Burkholderia species and the host complement system, with the goal of highlighting areas in this field that warrant further investigation.
... CihC, and lipopolysaccharide (27)(28)(29), respectively, to capture C1-INH on the cell surface, maintaining it in an active, inhibitory state. A hybrid of the above two strategies of C1-INH targeting has been proposed to be used by Escherichia coli O157:H7, involving capture of C1-INH on the cell surface followed by an enzymatic cleavage (30). While targeting an inhibitor to the pathogen surface is a self-evident way of enhancing immune evasion, the utility of destruction of C1-INH is less obvious, but it is explained by the fact that removal of C1-INH from serum leads to rapid, catastrophic activation of complement, leading to depletion of complement activity and so, perversely, less complement attack on the pathogen (22,25,26). ...
Complement, contact activation, coagulation, and fibrinolysis are serum protein cascades that need strict regulation to maintain human health. Serum glycoprotein, a C1 inhibitor (C1-INH), is a key regulator (inhibitor) of serine proteases of all the above-mentioned pathways. Recently, an autotransporter protein, virulence-associated gene 8 (Vag8), produced by the whooping cough pathogen, Bordetella pertussis , was shown to bind to C1-INH and interfere with its function. Here, we present the structure of the Vag8–C1-INH complex determined using cryo-electron microscopy at a 3.6-Å resolution. The structure shows a unique mechanism of C1-INH inhibition not employed by other pathogens, where Vag8 sequesters the reactive center loop of C1-INH, preventing its interaction with the target proteases.
IMPORTANCE The structure of a 10-kDa protein complex is one of the smallest to be determined using cryo-electron microscopy at high resolution. The structure reveals that C1-INH is sequestered in an inactivated state by burial of the reactive center loop in Vag8. By so doing, the bacterium is able to simultaneously perturb the many pathways regulated by C1-INH. Virulence mechanisms such as the one described here assume more importance given the emerging evidence about dysregulation of contact activation, coagulation, and fibrinolysis leading to COVID-19 pneumonia.
... The regulators of complement pathways C1-inhibitor (C1-INH) (Davis et al. 1986), C4b-binding protein (C4BP), MBL/ficolin/CLassociated protein-1 (MAP-1), and small MBL-associated protein (sMAP) (Schmidt et al. 2016) are utilized by various pathogens for their survival. Escherichia coli and Bordetella pertussis use C1-INH (Lathem et al. 2004;Marr et al. 2007), many pathogens exploit C4BP as part of their survival strategy Ram 2008, Hovingh et al. 2016). As coagulation inhibitors can affect LP, such as anti-thrombin inactivation of MASPs (Presanis et al. 2004), it may also be part of the survival strategy of various pathogens. ...
Mannan-binding lectin (MBL), a pathogen recognition receptor of the innate immune system, plays a key role in all types of infections and diseases. The ability of MBL to recognize pathogens facilitates immune mechanisms to function efficiently for the clearance of disease-causing agents. MBL not only initiates activation of lectin complement pathway but also simultaneously facilitates other effector functions of the immune system, such as proinflammatory responses, generation of reactive oxygen species, and phagocytosis. Thus, appropriate levels of MBL provide protection from the majority of diseases. However, certain intracellular organisms whose phagocytosis is increased due to opsonization with MBL get benefits from higher levels of MBL and thus in such cases MBL becomes a facilitator for these organisms to cause diseases. MBL being protective or increasing the susceptibility to various bacterial, viral, parasitic, and fungal infections has been extensively studied. Few important polymorphic sites in the exon and promoter region of MBL gene have been reported and they cause extensive variations in serum MBL concentration. Association of occurrence of diseases with the genetic variants has also been scientifically proven. Even though the immune system of the host is well equipped, pathogens have evolved to employ various survival strategies and evade immune response to cause disease.KeywordsMannan-Binding LectinMBL-associated serine protease MASPDisease association
