Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 143
Filtrar
Más filtros

Banco de datos
País/Región como asunto
Tipo del documento
Intervalo de año de publicación
1.
Annu Rev Microbiol ; 2024 Aug 15.
Artículo en Inglés | MEDLINE | ID: mdl-39146354

RESUMEN

Staphylococcus aureus is a commensal of the skin and nares of humans as well as the causative agent of infections associated with significant mortality. The acquisition of antibiotic resistance traits complicates the treatment of such infections and has prompted the development of monoclonal antibodies. The selection of protective antigens is typically guided by studying the natural antibody responses to a pathogen. What happens when the pathogen masks these antigens and subverts adaptive responses, or when the pathogen inhibits or alters the effector functions of antibodies? S. aureus is constantly exposed to its human host and has evolved all these strategies. Here, we review how anti-S. aureus targets have been selected and how antibodies have been engineered to overcome the formidable immune evasive activities of this pathogen. We discuss the prospects of antibody-based therapeutics in the context of disease severity, immune competence, and history of past infections.

2.
Nature ; 611(7935): 312-319, 2022 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-36261521

RESUMEN

Infectious diseases are among the strongest selective pressures driving human evolution1,2. This includes the single greatest mortality event in recorded history, the first outbreak of the second pandemic of plague, commonly called the Black Death, which was caused by the bacterium Yersinia pestis3. This pandemic devastated Afro-Eurasia, killing up to 30-50% of the population4. To identify loci that may have been under selection during the Black Death, we characterized genetic variation around immune-related genes from 206 ancient DNA extracts, stemming from two different European populations before, during and after the Black Death. Immune loci are strongly enriched for highly differentiated sites relative to a set of non-immune loci, suggesting positive selection. We identify 245 variants that are highly differentiated within the London dataset, four of which were replicated in an independent cohort from Denmark, and represent the strongest candidates for positive selection. The selected allele for one of these variants, rs2549794, is associated with the production of a full-length (versus truncated) ERAP2 transcript, variation in cytokine response to Y. pestis and increased ability to control intracellular Y. pestis in macrophages. Finally, we show that protective variants overlap with alleles that are today associated with increased susceptibility to autoimmune diseases, providing empirical evidence for the role played by past pandemics in shaping present-day susceptibility to disease.


Asunto(s)
ADN Antiguo , Predisposición Genética a la Enfermedad , Inmunidad , Peste , Selección Genética , Yersinia pestis , Humanos , Aminopeptidasas/genética , Aminopeptidasas/inmunología , Peste/genética , Peste/inmunología , Peste/microbiología , Peste/mortalidad , Yersinia pestis/inmunología , Yersinia pestis/patogenicidad , Selección Genética/inmunología , Europa (Continente)/epidemiología , Europa (Continente)/etnología , Inmunidad/genética , Conjuntos de Datos como Asunto , Londres/epidemiología , Dinamarca/epidemiología
3.
PLoS Pathog ; 20(6): e1012306, 2024 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-38843309

RESUMEN

Staphylococcus aureus is a human-adapted pathogen that replicates by asymptomatically colonizing its host. S. aureus is also the causative agent of purulent skin and soft tissue infections as well as bloodstream infections that result in the metastatic seeding of abscess lesions in all organ tissues. Prolonged colonization, infection, disease relapse, and recurrence point to the versatile capacity of S. aureus to bypass innate and adaptive immune defenses as well as the notion that some hosts fail to generate protective immune responses. Here, we find a genetic trait that provides protection against this pathogen. Mice lacking functional H2-O, the equivalent of human HLA-DO, inoculated with a mouse-adapted strain of S. aureus, efficiently decolonize the pathogen. Further, these decolonized animals resist subsequent bloodstream challenge with methicillin-resistant S. aureus. A genetic approach demonstrates that T-cell dependent B cell responses are required to control S. aureus colonization and infection in H2-O-deficient mice. Reduced bacterial burdens in these animals correlate with increased titers and enhanced phagocytic activity of S. aureus-specific antibodies. H2-O negatively regulates the loading of high affinity peptides on major histocompatibility class II (MHC-II) molecules. Thus, we hypothesize that immune responses against S. aureus are derepressed in mice lacking H2-O because more high affinity peptides are presented by MHC-II. We speculate that loss-of-function HLA-DO alleles may similarly control S. aureus replication in humans.


Asunto(s)
Infecciones Estafilocócicas , Staphylococcus aureus , Animales , Infecciones Estafilocócicas/inmunología , Infecciones Estafilocócicas/microbiología , Ratones , Staphylococcus aureus/inmunología , Ratones Noqueados , Ratones Endogámicos C57BL , Antígenos de Histocompatibilidad Clase II/inmunología , Staphylococcus aureus Resistente a Meticilina/inmunología , Humanos
4.
Proc Natl Acad Sci U S A ; 120(14): e2220765120, 2023 04 04.
Artículo en Inglés | MEDLINE | ID: mdl-36972444

RESUMEN

Antibodies bind target molecules with exquisite specificity. The removal of these targets is mediated by the effector functions of antibodies. We reported earlier that the monoclonal antibody (mAb) 3F6 promotes opsonophagocytic killing of Staphylococcus aureus in blood and reduces bacterial replication in animals. Here, we generated mouse immunoglobulin G (mIgG) subclass variants and observed a hierarchy in protective efficacy 3F6-mIgG2a > 3F6-mIgG1 ≥ 3F6-mIgG2b >> 3F6-mIgG3 following bloodstream challenge of C57BL/6J mice. This hierarchy was not observed in BALB/cJ mice: All IgG subclasses conferred similar protection. IgG subclasses differ in their ability to activate complement and interact with Fcγ receptors (FcγR) on immune cells. 3F6-mIgG2a-dependent protection was lost in FcγR-deficient, but not in complement-deficient C57BL/6J animals. Measurements of the relative ratio of FcγRIV over complement receptor 3 (CR3) on neutrophils suggest the preferential expression of FcγRIV in C57BL/6 mice and of CR3 in BALB/cJ mice. To determine the physiological significance of these differing ratios, blocking antibodies against FcγRIV or CR3 were administered to animals before challenge. Correlating with the relative abundance of each receptor, 3F6-mIgG2a-dependent protection in C57BL/6J mice showed a greater reliance for FcγRIV while protection in BALB/cJ mice was only impaired upon neutralization of CR3. Thus, 3F6-based clearance of S. aureus in mice relies on a strain-specific contribution of variable FcγR- and complement-dependent pathways. We surmise that these variabilities are the result of genetic polymorphism(s) that may be encountered in other mammals including humans and may have clinical implications in predicting the efficacy of mAb-based therapies.


Asunto(s)
Inmunoglobulina G , Staphylococcus aureus , Humanos , Ratones , Animales , Staphylococcus aureus/metabolismo , Receptores de IgG/genética , Ratones Endogámicos C57BL , Anticuerpos Monoclonales/farmacología , Proteínas del Sistema Complemento , Mamíferos/metabolismo
5.
Nature ; 574(7776): 57-62, 2019 10.
Artículo en Inglés | MEDLINE | ID: mdl-31534221

RESUMEN

The causative agent of plague, Yersinia pestis, uses a type III secretion system to selectively destroy immune cells in humans, thus enabling Y. pestis to reproduce in the bloodstream and be transmitted to new hosts through fleabites. The host factors that are responsible for the selective destruction of immune cells by plague bacteria are unknown. Here we show that LcrV, the needle cap protein of the Y. pestis type III secretion system, binds to the N-formylpeptide receptor (FPR1) on human immune cells to promote the translocation of bacterial effectors. Plague infection in mice is characterized by high mortality; however, Fpr1-deficient mice have increased survival and antibody responses that are protective against plague. We identified FPR1R190W as a candidate resistance allele in humans that protects neutrophils from destruction by the Y. pestis type III secretion system. Thus, FPR1 is a plague receptor on immune cells in both humans and mice, and its absence or mutation provides protection against Y. pestis. Furthermore, plague selection of FPR1 alleles appears to have shaped human immune responses towards other infectious diseases and malignant neoplasms.


Asunto(s)
Macrófagos/metabolismo , Neutrófilos/metabolismo , Peste/microbiología , Receptores de Formil Péptido/metabolismo , Yersinia pestis/metabolismo , Alelos , Animales , Antígenos Bacterianos/metabolismo , Adhesión Bacteriana , Sistemas CRISPR-Cas , Quimiotaxis/inmunología , Modelos Animales de Enfermedad , Femenino , Células HEK293 , Humanos , Macrófagos/citología , Macrófagos/inmunología , Macrófagos/microbiología , Masculino , Ratones , Ratones Endogámicos C57BL , Neutrófilos/citología , Neutrófilos/inmunología , Neutrófilos/microbiología , Peste/inmunología , Peste/prevención & control , Polimorfismo de Nucleótido Simple/genética , Proteínas Citotóxicas Formadoras de Poros/metabolismo , Receptores de Formil Péptido/antagonistas & inhibidores , Receptores de Formil Péptido/deficiencia , Receptores de Formil Péptido/genética , Sistemas de Secreción Tipo III/efectos de los fármacos , Células U937 , Yersinia pestis/química , Yersinia pestis/inmunología , Yersinia pestis/patogenicidad
6.
Proc Natl Acad Sci U S A ; 119(4)2022 01 25.
Artículo en Inglés | MEDLINE | ID: mdl-35058363

RESUMEN

Gram-positive organisms with their thick envelope cannot be lysed by complement alone. Nonetheless, antibody-binding on the surface can recruit complement and mark these invaders for uptake and killing by phagocytes, a process known as opsonophagocytosis. The crystallizable fragment of immunoglobulins (Fcγ) is key for complement recruitment. The cell surface of S. aureus is coated with Staphylococcal protein A (SpA). SpA captures the Fcγ domain of IgG and interferes with opsonization by anti-S. aureus antibodies. In principle, the Fcγ domain of therapeutic antibodies could be engineered to avoid the inhibitory activity of SpA. However, the SpA-binding site on Fcγ overlaps with that of the neonatal Fc receptor (FcRn), an interaction that is critical for prolonging the half-life of serum IgG. This evolutionary adaptation poses a challenge for the exploration of Fcγ mutants that can both weaken SpA-IgG interactions and retain stability. Here, we use both wild-type and transgenic human FcRn mice to identify antibodies with enhanced half-life and increased opsonophagocytic killing in models of S. aureus infection and demonstrate that antibody-based immunotherapy can be improved by modifying Fcγ. Our experiments also show that by competing for FcRn-binding, staphylococci effectively reduce the half-life of antibodies during infection. These observations may have profound impact in treating cancer, autoimmune, and asthma patients colonized or infected with S. aureus and undergoing monoclonal antibody treatment.


Asunto(s)
Anticuerpos Antibacterianos/genética , Anticuerpos Antibacterianos/inmunología , Anticuerpos Monoclonales/genética , Anticuerpos Monoclonales/inmunología , Opsonización/inmunología , Ingeniería de Proteínas , Secuencia de Aminoácidos , Citotoxicidad Celular Dependiente de Anticuerpos/inmunología , Activación de Complemento , Relación Dosis-Respuesta a Droga , Relación Dosis-Respuesta Inmunológica , Humanos , Fagocitosis/inmunología , Unión Proteica , Ingeniería de Proteínas/métodos , Dominios y Motivos de Interacción de Proteínas/genética , Dominios y Motivos de Interacción de Proteínas/inmunología , Receptores Fc/genética , Proteína Estafilocócica A/inmunología , Staphylococcus aureus/inmunología
7.
Proc Natl Acad Sci U S A ; 119(14): e2119093119, 2022 04 05.
Artículo en Inglés | MEDLINE | ID: mdl-35312341

RESUMEN

SignificanceUsing SARS-CoV-2 as a relevant case study for infectious disease, we investigate the structure-function relationships that dictate antiviral spherical nucleic acid (SNA) vaccine efficacy. We show that the SNA architecture can be rapidly employed to target COVID-19 through incorporation of the receptor-binding domain, and that the resulting vaccine potently activates human cells in vitro and mice in vivo. Furthermore, when challenged with a lethal viral infection, only mice treated with the SNA vaccine survived. Taken together, this work underscores the importance of rational vaccine design for infectious disease to yield vaccines that elicit more potent immune responses to effectively fight disease.


Asunto(s)
Control de Enfermedades Transmisibles , Ácidos Nucleicos/inmunología , Vacunas de ADN/inmunología , Animales , Biotecnología , COVID-19/prevención & control , Control de Enfermedades Transmisibles/métodos , Enfermedades Transmisibles/etiología , Enfermedades Transmisibles/inmunología , Humanos , Ácidos Nucleicos/química , SARS-CoV-2/inmunología , Desarrollo de Vacunas , Vacunas de ADN/genética , Vacunas Virales/genética , Vacunas Virales/inmunología
8.
Arterioscler Thromb Vasc Biol ; 43(2): 267-285, 2023 02.
Artículo en Inglés | MEDLINE | ID: mdl-36453281

RESUMEN

BACKGROUND: Infective endocarditis (IE) is characterized by an infected thrombus at the heart valves. How bacteria bypass the immune system and cause these thrombi remains unclear. Neutrophils releasing NETs (neutrophil extracellular traps) lie at this interface between host defense and coagulation. We aimed to determine the role of NETs in IE immunothrombosis. METHODS: We used a murine model of Staphylococcus aureus endocarditis in which IE is provoked on inflamed heart valves and characterized IE thrombus content by immunostaining identifying NETs. Antibody-mediated neutrophil depletion and neutrophil-selective PAD4 (peptidylarginine deiminase 4)-knockout mice were used to clarify the role of neutrophils and NETs, respectively. S. aureus mutants deficient in key virulence factors related to immunothrombosis (nucleases or staphylocoagulases) were investigated. RESULTS: Neutrophils releasing NETs were present in infected thrombi and within cellular infiltrates in the surrounding vasculature. Neutrophil depletion increased occurrence of IE, whereas neutrophil-selective impairment of NET formation did not alter IE occurrence. Absence of S. aureus nuclease, which degrades NETs, did not affect endocarditis outcome. In contrast, absence of staphylocoagulases (coagulase and von Willebrand factor binding protein) led to improved survival, decreased bacteremia, smaller infiltrates, and decreased tissue destruction. Significantly more NETs were present in these vegetations, which correlated with decreased bacteria and cell death in the adjacent vascular wall. CONCLUSIONS: Neutrophils protect against IE independent of NET release. Absence of S. aureus coagulases, but not nucleases, reduced IE severity and increased NET levels. Staphylocoagulase-induced fibrin likely hampers NETs from constraining infection and the resultant tissue damage, a hallmark of valve destruction in IE.


Asunto(s)
Endocarditis Bacteriana , Endocarditis , Trampas Extracelulares , Infecciones Estafilocócicas , Ratones , Animales , Neutrófilos/metabolismo , Trampas Extracelulares/metabolismo , Staphylococcus aureus , Tromboinflamación , Endocarditis Bacteriana/prevención & control , Endocarditis Bacteriana/metabolismo , Endocarditis/metabolismo
9.
J Biol Chem ; 298(9): 102318, 2022 09.
Artículo en Inglés | MEDLINE | ID: mdl-35921891

RESUMEN

Secretion systems utilize ATPase activity to facilitate the translocation of proteins into and across membranes. In bacteria, the universally conserved SecA ATPase binds a large repertoire of preproteins and interacts with the SecYEG translocon. In contrast, the type 7b secretion system (T7bSS) of Staphylococcus aureus supports the secretion of a restricted subset of proteins. T7bSSs are found in several Firmicutes as gene clusters encoding secreted WXG100 proteins and FtsK/SpoIIIE-like ATPase. In S. aureus, this ATPase is called EssC and comprises two cytosolic forkhead-associated domains (FHA1-2), two membrane-spanning segments (TM1-2), and four cytosolic modules named DUF (domain of unknown function) and ATPases1-3 (D1D2D3). However, a detailed understanding of the interactions of EssC in the T7bSS is not clear. Here, we tagged EssC and performed affinity chromatography of detergent-solubilized extracts of wild type and isogenic mutants of S. aureus. We found that EssC recruits EsaA, EssA, and EssB in a complex referred to as the ESS (ESAT-6 like secretion system) translocon, and secreted substrates were not required for translocon assembly. Furthermore, deletions of FHA1 and DUF rendered EssC unstable, whereas FHA2 was required for association with EssB. This interaction was independent of EsaA, but EsaA was required to recruit EssA to the EssC-EssB complex. Finally, we show that assembly of the ESS translocon was impaired upon mutation of D2 structural motifs. Together, our data indicate that the ESS translocon is maintained fully assembled at the plasma membrane and that D2 is fundamental in sustaining the integrity of this complex.


Asunto(s)
Adenosina Trifosfatasas , Proteínas Bacterianas , Staphylococcus aureus , Sistemas de Secreción Tipo VII , Adenosina Trifosfatasas/genética , Adenosina Trifosfatasas/metabolismo , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Transporte de Proteínas , Canales de Translocación SEC/genética , Canales de Translocación SEC/metabolismo , Staphylococcus aureus/genética , Staphylococcus aureus/metabolismo , Sistemas de Secreción Tipo VII/metabolismo
10.
Infect Immun ; 91(5): e0001523, 2023 05 16.
Artículo en Inglés | MEDLINE | ID: mdl-37039657

RESUMEN

Staphylococcus aureus bears a type 7b secretion system (T7bSS) that assembles in the bacterial envelope to promote the secretion of WXG-like proteins and toxic effectors bearing LXG domains. Cognate immunity proteins bind cytosolic effectors to mute their toxicity prior to secretion. T7b-secreted factors have been associated with the pathogenesis of staphylococcal disease and intraspecies competition. We identified earlier strain WU1, an S. aureus ST88 isolate that caused outbreaks of skin and soft tissue infections in mouse breeding facilities. WU1 was also found to persistently colonize the nasopharynx of animals, suggesting a strong host adaptation. In this manner, WU1 colonization and infectivity in mice resembles that of methicillin-sensitive and -resistant S. aureus strains in humans, where nasal carriage is a major risk factor for invasive infections. Here, animals were colonized with wild-type or T7-deficient WU1 strains or combinations thereof. Absence of the T7bSS did not affect colonization in the nasopharynx of animals, and although fluctuations were observed in weekly samplings, the wild-type strain did not replace the T7-deficient strain in cocolonization experiments. Bloodstream infection with a T7b-deficient strain resulted in enhanced survival and reduced bacterial loads and abscesses in soft tissues compared to infection with wild-type WU1. Together, experiments using a mouse-adapted strain suggest that the T7bSS of S. aureus is an important contributor to the pathogenesis of invasive disease.


Asunto(s)
Staphylococcus aureus Resistente a Meticilina , Sepsis , Infecciones Estafilocócicas , Humanos , Animales , Staphylococcus aureus , Infecciones Estafilocócicas/microbiología , Piel
11.
Proc Natl Acad Sci U S A ; 117(37): 22992-23000, 2020 09 15.
Artículo en Inglés | MEDLINE | ID: mdl-32855300

RESUMEN

Antibodies may bind to bacterial pathogens or their toxins to control infections, and their effector activity is mediated through the recruitment of complement component C1q or the engagement with Fcγ receptors (FcγRs). For bacterial pathogens that rely on a single toxin to cause disease, immunity correlates with toxin neutralization. Most other bacterial pathogens, including Staphylococcus aureus, secrete numerous toxins and evolved multiple mechanisms to escape opsonization and complement killing. Several vaccine candidates targeting defined surface antigens of S. aureus have failed to meet clinical endpoints. It is unclear that such failures can be solely attributed to the poor selection of antibody targets. Thus far, studies to delineate antibody-mediated uptake and killing of Gram-positive pathogens remain extremely limited. Here, we exploit 3F6-hIgG1, a human monoclonal antibody that binds and neutralizes the abundant surface-exposed Staphylococcal protein A (SpA). We find that galactosylation of 3F6-hIgG1 that favors C1q recruitment is indispensable for opsonophagocytic killing of staphylococci and for protection against bloodstream infection in animals. However, the simple removal of fucosyl residues, which results in reduced C1q binding and increased engagement with FcγR, maintains the opsonophagocytic killing and protective attributes of the antibody. We confirm these results by engineering 3F6-hIgG1 variants with biased binding toward C1q or FcγRs. While the therapeutic benefit of monoclonal antibodies against infectious disease agents may be debatable, the functional characterization of such antibodies represents a powerful tool for the development of correlates of protection that may guide future vaccine trials.


Asunto(s)
Anticuerpos Antibacterianos/inmunología , Anticuerpos Monoclonales/inmunología , Fagocitosis/inmunología , Proteína Estafilocócica A/inmunología , Animales , Línea Celular , Glicosilación , Células HEK293 , Humanos , Ratones , Ratones Endogámicos BALB C , Infecciones Estafilocócicas/inmunología , Staphylococcus aureus/inmunología
12.
J Am Soc Nephrol ; 33(7): 1293-1307, 2022 07.
Artículo en Inglés | MEDLINE | ID: mdl-35236774

RESUMEN

BACKGROUND: Severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2) uses full-length angiotensin converting enzyme 2 (ACE2) as a main receptor to enter target cells. The goal of this study was to demonstrate the preclinical efficacy of a novel soluble ACE2 protein with increased duration of action and binding capacity in a lethal mouse model of COVID-19. METHODS: A human soluble ACE2 variant fused with an albumin binding domain (ABD) was linked via a dimerization motif hinge-like 4-cysteine dodecapeptide (DDC) to improve binding capacity to SARS-CoV-2. This novel soluble ACE2 protein (ACE2-1-618-DDC-ABD) was then administered intranasally and intraperitoneally to mice before intranasal inoculation of SARS-CoV-2 and then for two additional days post viral inoculation. RESULTS: Untreated animals became severely ill, and all had to be humanely euthanized by day 6 or 7 and had pulmonary alveolar hemorrhage with mononuclear infiltrates. In contrast, all but one mouse infected with a lethal dose of SARS-CoV-2 that received ACE2-1-618-DDC-ABD survived. In the animals inoculated with SARS-CoV-2 that were untreated, viral titers were high in the lungs and brain, but viral titers were absent in the kidneys. Some untreated animals, however, had variable degrees of kidney proximal tubular injury as shown by attenuation of the proximal tubular brush border and increased NGAL and TUNEL staining. Viral titers in the lung and brain were reduced or nondetectable in mice that received ACE2-1-618-DDC-ABD, and the animals developed only moderate disease as assessed by a near-normal clinical score, minimal weight loss, and improved lung and kidney injury. CONCLUSIONS: This study demonstrates the preclinical efficacy of a novel soluble ACE2 protein, termed ACE2-1-618-DDC-ABD, in a lethal mouse model of SARS-CoV-2 infection that develops severe lung injury and variable degrees of moderate kidney proximal tubular injury.


Asunto(s)
Enzima Convertidora de Angiotensina 2 , COVID-19 , Enzima Convertidora de Angiotensina 2/uso terapéutico , Animales , COVID-19/terapia , Riñón/virología , Pulmón/virología , Ratones , SARS-CoV-2
13.
J Bacteriol ; 204(9): e0025322, 2022 Sep 20.
Artículo en Inglés | MEDLINE | ID: mdl-35997505

RESUMEN

Bacillus anthracis elaborates a secondary cell wall polysaccharide (SCWP) made of 6 to 12 trisaccharide units. Pyruvyl and acetyl substitutions of the distal unit are prerequisites for the noncovalent retention of 22 secreted Bacillus S-layer (Bsl)-associated proteins bearing an S-layer homology (SLH) domain. Surface display of Bsl proteins contributes to cell separation as well as virulence. Earlier work suggested that TagO initiates the synthesis of SCWP while GneY and GneZ, two UDP-GlcNAc 2-epimerases, synthesize ManNAc that is later incorporated in the repeat unit (→4)-ManNAc-(ß1→4)-GlcNAc-(ß1→6)-GlcNAc-(α1→). In organisms that synthesize wall teichoic acid, TagA catalysts have been shown to form the glycosidic bond ManNAc-(ß1→4)-GlcNAc. Here, we show that genes bas2675 and bas5272, predicted to encode glycosyltransferases of the WecB/TagA/CpsF family (PFAM03808; CAZy GT26), are required for B. anthracis SCWP synthesis and S-layer assembly. Similar to tagO or gneY gneZ mutants, B. anthracis strains depleted of tagA1 (bas5272) cannot maintain cell shape, support vegetative growth, or synthesize SCWP. Expression of tagA2 (bas2675), or Staphylococcus aureus tagA on a plasmid, rescues the nonviable tagA1 mutant. We propose that TagA1 and TagA2 fulfill overlapping and key glycosyltransferase functions for the synthesis of repeat units of the SCWP of B. anthracis. IMPORTANCE Glycosyltransferases (GTs) catalyze the transfer of sugar moieties from activated donor molecules to acceptor molecules to form glycosidic bonds using a retaining or inverting mechanism. Based on the structural relatedness of their catalytic and carbohydrate-binding modules, GTs have been grouped into 115 families in the Carbohydrate-Active EnZyme (CAZy) database. For complex products, the functional assignment of GTs remains highly challenging without the knowledge of the chemical structure of the assembled polymer. Here, we propose that two uncharacterized GTs of B. anthracis belonging to the WecB/TagA/CpsF family incorporate ManNAc in repeat units of the secondary cell wall polymer of bacilli species.


Asunto(s)
Bacillus anthracis , Bacillus anthracis/metabolismo , Proteínas Bacterianas/metabolismo , Pared Celular/metabolismo , Glicosiltransferasas/genética , Glicosiltransferasas/metabolismo , Glicoproteínas de Membrana/metabolismo , Polímeros , Polisacáridos/metabolismo , Azúcares/metabolismo , Trisacáridos/química , Uridina Difosfato/análisis , Uridina Difosfato/metabolismo
14.
Annu Rev Microbiol ; 71: 79-98, 2017 09 08.
Artículo en Inglés | MEDLINE | ID: mdl-28622090

RESUMEN

Bacillus anthracis, the anthrax agent, is a member of the Bacillus cereus sensu lato group, which includes invasive pathogens of mammals or insects as well as nonpathogenic environmental strains. The genes for anthrax pathogenesis are located on two large virulence plasmids. Similar virulence plasmids have been acquired by other B. cereus strains and enable the pathogenesis of anthrax-like diseases. Among the virulence factors of B. anthracis is the S-layer-associated protein BslA, which endows bacilli with invasive attributes for mammalian hosts. BslA surface display and function are dependent on the bacterial S-layer, whose constituents assemble by binding to the secondary cell wall polysaccharide (SCWP) via S-layer homology (SLH) domains. B. anthracis and other pathogenic B. cereus isolates harbor genes for the secretion of S-layer proteins, for S-layer assembly, and for synthesis of the SCWP. We review here recent insights into the assembly and function of the S-layer and the SCWP.


Asunto(s)
Bacillus anthracis/metabolismo , Glicoproteínas de Membrana/metabolismo , Multimerización de Proteína
15.
Proc Natl Acad Sci U S A ; 116(39): 19659-19664, 2019 09 24.
Artículo en Inglés | MEDLINE | ID: mdl-31413191

RESUMEN

Rickettsial diseases have long been diagnosed with serum antibodies cross-reactive against Proteus vulgaris (Weil-Felix reaction). Although Weil-Felix antibodies are associated with the development of immunity, their rickettsial target and contribution to disease pathogenesis are not established. Here, we developed a transposon for insertional mutagenesis of Rickettsia conorii, isolating variants defective for replication in cultured cells and in spotted fever pathogenesis. Mutations in the polysaccharide synthesis operon (pso) abolish lipopolysaccharide O-antigen synthesis and Weil-Felix serology and alter outer-membrane protein assembly. Unlike wild-type R. conorii, pso mutants cannot elicit bactericidal antibodies that bind O antigen. The pso operon is conserved among rickettsial pathogens, suggesting that bactericidal antibodies targeting O antigen may generate universal immunity that could be exploited to develop vaccines against rickettsial diseases.


Asunto(s)
Reacciones Cruzadas/inmunología , Antígenos O/inmunología , Rickettsia conorii/inmunología , Antibacterianos , Anticuerpos Antibacterianos/inmunología , Lipopolisacáridos/inmunología , Rickettsia/inmunología , Rickettsia/patogenicidad , Infecciones por Rickettsia/inmunología , Rickettsia conorii/patogenicidad
16.
J Bacteriol ; 203(9)2021 05 01.
Artículo en Inglés | MEDLINE | ID: mdl-33593946

RESUMEN

LyrA (SpdC), a homologue of eukaryotic CAAX proteases that act on prenylated substrates, has been implicated in the assembly of several pathways of the envelope of Staphylococcus aureus. We described earlier the Lysostaphin resistance (Lyr) and Staphylococcal protein A display (Spd) phenotypes associated with loss of the lyrA (spdC) gene. However, a direct contribution to the assembly of pentaglycine crossbridges, the target of lysostaphin cleavage in S. aureus peptidoglycan, or of Staphylococcal protein A attachment to peptidoglycan could not be attributed directly to LyrA (SpdC). These two processes are catalyzed by the Fem factors and Sortase A, respectively. To gain insight into the function of LyrA (SpdC), here we use affinity chromatography and LC-MS/MS analysis and report that LyrA interacts with SagB. SagB cleaves glycan strands of peptidoglycan to achieve physiological length. Similar to sagB peptidoglycan, lyrA peptidoglycan contains extended glycan strands. Purified lyrA peptidoglycan can still be cleaved to physiological length by SagB in vitro LyrA does not modify or cleave peptidoglycan, it also does not modify or stabilize SagB. The membrane bound domain of LyrA is sufficient to support SagB activity but predicted 'CAAX enzyme' catalytic residues in this domain are dispensable. We speculate that LyrA exerts its effect on bacterial prenyl substrates, specifically undecaprenol-bound peptidoglycan substrates of SagB, to help control glycan length. Such an activity also explains the Lyr and Spd phenotypes observed earlier.IMPORTANCE Peptidoglycan is assembled on the trans side of the plasma membrane from lipid II precursors into glycan chains that are crosslinked at stem peptides. In S. aureus, SagB, a membrane-associated N-acetylglucosaminidase, cleaves polymerized glycan chains to their physiological length. Deletion of sagB is associated with longer glycan strands in peptidoglycan, altered protein trafficking and secretion in the envelope, and aberrant excretion of cytosolic proteins. It is not clear whether SagB, with its single transmembrane segment, serves as the molecular ruler of glycan chains or whether other factors modulate its activity. Here, we show that LyrA (SpdC), a protein of the CAAX type II prenyl endopeptidase family, modulates SagB activity via interaction though its transmembrane domain.

17.
J Biol Chem ; 295(39): 13664-13676, 2020 09 25.
Artículo en Inglés | MEDLINE | ID: mdl-32759309

RESUMEN

Staphylococcal peptidoglycan is characterized by pentaglycine cross-bridges that are cross-linked between adjacent wall peptides by penicillin-binding proteins to confer robustness and flexibility. In Staphylococcus aureus, pentaglycine cross-bridges are synthesized by three proteins: FemX adds the first glycine, and the homodimers FemA and FemB sequentially add two Gly-Gly dipeptides. Occasionally, serine residues are also incorporated into the cross-bridges by enzymes that have heretofore not been identified. Here, we show that the FemA/FemB homologues FmhA and FmhC pair with FemA and FemB to incorporate Gly-Ser dipeptides into cross-bridges and to confer resistance to lysostaphin, a secreted bacteriocin that cleaves the pentaglycine cross-bridge. FmhA incorporates serine residues at positions 3 and 5 of the cross-bridge. In contrast, FmhC incorporates a single serine at position 5. Serine incorporation also lowers resistance toward oxacillin, an antibiotic that targets penicillin-binding proteins, in both methicillin-sensitive and methicillin-resistant strains of S. aureus FmhC is encoded by a gene immediately adjacent to lytN, which specifies a hydrolase that cleaves the bond between the fifth glycine of cross-bridges and the alanine of the adjacent stem peptide. In this manner, LytN facilitates the separation of daughter cells. Cell wall damage induced upon lytN overexpression can be alleviated by overexpression of fmhC. Together, these observations suggest that FmhA and FmhC generate peptidoglycan cross-bridges with unique serine patterns that provide protection from endogenous murein hydrolases governing cell division and from bacteriocins produced by microbial competitors.


Asunto(s)
Proteínas Bacterianas/metabolismo , Peptidoglicano/metabolismo , Serina/metabolismo , Staphylococcus aureus/metabolismo
18.
Proc Natl Acad Sci U S A ; 115(26): 6846-6851, 2018 06 26.
Artículo en Inglés | MEDLINE | ID: mdl-29891696

RESUMEN

Staphylococcus aureus colonizes large segments of the human population and causes invasive infections due to its ability to escape phagocytic clearance. During infection, staphylococcal nuclease and adenosine synthase A convert neutrophil extracellular traps to deoxyadenosine (dAdo), which kills phagocytes. The mechanism whereby staphylococcal dAdo intoxicates phagocytes is not known. Here we used CRISPR-Cas9 mutagenesis to show that phagocyte intoxication involves uptake of dAdo via the human equilibrative nucleoside transporter 1, dAdo conversion to dAMP by deoxycytidine kinase and adenosine kinase, and signaling via subsequent dATP formation to activate caspase-3-induced cell death. Disruption of this signaling cascade confers resistance to dAdo-induced intoxication of phagocytes and may provide therapeutic opportunities for the treatment of infections caused by antibiotic-resistant S. aureus strains.


Asunto(s)
Caspasa 3/metabolismo , Desoxiadenosinas/metabolismo , Trampas Extracelulares/metabolismo , Neutrófilos/metabolismo , Infecciones Estafilocócicas/metabolismo , Staphylococcus aureus/metabolismo , Caspasa 3/genética , Muerte Celular/genética , Desoxiadenosinas/genética , Trampas Extracelulares/genética , Trampas Extracelulares/microbiología , Humanos , Neutrófilos/microbiología , Neutrófilos/patología , Infecciones Estafilocócicas/genética , Infecciones Estafilocócicas/patología , Staphylococcus aureus/genética
19.
J Bacteriol ; 202(15)2020 07 09.
Artículo en Inglés | MEDLINE | ID: mdl-32457049

RESUMEN

Bacillus anthracis, the causative agent of anthrax disease, elaborates a secondary cell wall polysaccharide (SCWP) that is required for the retention of surface layer (S-layer) and S-layer homology (SLH) domain proteins. Genetic disruption of the SCWP biosynthetic pathway impairs growth and cell division. B. anthracis SCWP is comprised of trisaccharide repeats composed of one ManNAc and two GlcNAc residues with O-3-α-Gal and O-4-ß-Gal substitutions. UDP-Gal, synthesized by GalE1, is the substrate of galactosyltransferases that modify the SCWP repeat. Here, we show that the gtsE gene, which encodes a predicted glycosyltransferase with a GT-A fold, is required for O-4-ß-Gal modification of trisaccharide repeats. We identify a DXD motif critical for GtsE activity. Three distinct genes, gtsA, gtsB, and gtsC, are required for O-3-α-Gal modification of trisaccharide repeats. Based on the similarity with other three-component glycosyltransferase systems, we propose that GtsA transfers Gal from cytosolic UDP-Gal to undecaprenyl phosphate (C55-P), GtsB flips the C55-P-Gal intermediate to the trans side of the membrane, and GtsC transfers Gal onto trisaccharide repeats. The deletion of galE1 does not affect growth in vitro, suggesting that galactosyl modifications are dispensable for the function of SCWP. The deletion of gtsA, gtsB, or gtsC leads to a loss of viability, yet gtsA and gtsC can be deleted in strains lacking galE1 or gtsE We propose that the loss of viability is caused by the accumulation of undecaprenol-bound precursors and present an updated model for SCWP assembly in B. anthracis to account for the galactosylation of repeat units.IMPORTANCE Peptidoglycan is a conserved extracellular macromolecule that protects bacterial cells from turgor pressure. Peptidoglycan of Gram-positive bacteria serves as a scaffold for the attachment of polymers that provide defined bacterial interactions with their environment. One such polymer, B. anthracis SCWP, is pyruvylated at its distal end to serve as a receptor for secreted proteins bearing the S-layer homology domain. Repeat units of SCWP carry three galactoses in B. anthracis Glycosylation is a recurring theme in nature and often represents a means to mask or alter conserved molecular signatures from intruders such as bacteriophages. Several glycosyltransferase families have been described based on bioinformatics prediction, but few have been studied. Here, we describe the glycosyltransferases that mediate the galactosylation of B. anthracis SCWP.


Asunto(s)
Bacillus anthracis/metabolismo , Pared Celular/metabolismo , Galactosa/metabolismo , Polisacáridos Bacterianos/metabolismo , Carbunco/microbiología , Bacillus anthracis/genética , Bacillus anthracis/crecimiento & desarrollo , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Pared Celular/química , Pared Celular/genética , Eliminación de Gen , Regulación Bacteriana de la Expresión Génica , Glicosilación , Humanos , Polisacáridos Bacterianos/química
20.
Eur Heart J ; 40(39): 3248-3259, 2019 10 14.
Artículo en Inglés | MEDLINE | ID: mdl-30945735

RESUMEN

AIMS: The pathogenesis of endocarditis is not well understood resulting in unsuccessful attempts at prevention. Clinical observations suggest that Staphylococcus aureus infects either damaged or inflamed heart valves. Using a newly developed endocarditis mouse model, we therefore studied the initial adhesion of S. aureus in both risk states. METHODS AND RESULTS: Using 3D confocal microscopy, we examined the adhesion of fluorescent S. aureus to murine aortic valves. To mimic different risk states we either damaged the valves with a surgically placed catheter or simulated valve inflammation by local endothelium activation. We used von Willebrand factor (VWF) gene-deficient mice, induced platelet and fibrinogen depletion and used several S. aureus mutant strains to investigate the contribution of both host and bacterial factors in early bacterial adhesion. Both cardiac valve damage and inflammation predisposed to endocarditis, but by distinct mechanisms. Following valve damage, S. aureus adhered directly to VWF and fibrin, deposited on the damaged valve. This was mediated by Sortase A-dependent adhesins such as VWF-binding protein and Clumping factor A. Platelets did not contribute. In contrast, upon cardiac valve inflammation, widespread endothelial activation led to endothelial cell-bound VWF release. This recruited large amounts of platelets, capturing S. aureus to the valve surface. Here, neither fibrinogen, nor Sortase A were essential. CONCLUSION: Cardiac valve damage and inflammation predispose to S. aureus endocarditis via distinct mechanisms. These findings may have important implications for the development of new preventive strategies, as some interventions might be effective in one risk state, but not in the other.


Asunto(s)
Válvula Aórtica/microbiología , Adhesión Bacteriana , Endocarditis Bacteriana/microbiología , Inflamación/complicaciones , Infecciones Estafilocócicas/complicaciones , Staphylococcus aureus/fisiología , Animales , Válvula Aórtica/lesiones , Plaquetas , Coagulasa/metabolismo , Modelos Animales de Enfermedad , Endocarditis Bacteriana/metabolismo , Endotelio/metabolismo , Femenino , Fibrina/metabolismo , Inflamación/metabolismo , Masculino , Ratones , Glicoproteínas de Membrana Plaquetaria/metabolismo , Infecciones Estafilocócicas/metabolismo , Staphylococcus aureus/metabolismo , Factor de von Willebrand/genética , Factor de von Willebrand/metabolismo
SELECCIÓN DE REFERENCIAS
DETALLE DE LA BÚSQUEDA