RESUMEN

The soil-dwelling bacterium Listeria monocytogenes survives a multitude of conditions when residing in the outside environment and as a pathogen within host cells. Key to survival within the infected mammalian host is the expression of bacterial gene products necessary for nutrient acquisition. Similar to many bacteria, L. monocytogenes uses peptide import to acquire amino acids. Peptide transport systems play an important role in nutrient uptake as well as in additional functions that include bacterial quorum sensing and signal transduction, recycling of peptidoglycan fragments, adherence to eukaryotic cells, and alterations in antibiotic susceptibility. It has been previously described that CtaP, encoded by lmo0135, is a multifunctional protein associated with activities that include cysteine transport, resistance to acid, membrane integrity, and bacterial adherence to host cells. ctaP is located next to two genes predicted to encode membrane-bound permeases lmo0136 and lmo0137, termed CtpP1 and CtpP2, respectively. Here, we show that CtpP1 and CtpP2 are required for bacterial growth in the presence of low concentrations of cysteine and for virulence in mouse infection models. Taken together, the data identify distinct nonoverlapping roles for two related permeases that are important for the growth and survival of L. monocytogenes within host cells. IMPORTANCE Bacterial peptide transport systems are important for nutrient uptake and may additionally function in a variety of other roles, including bacterial communication, signal transduction, and bacterial adherence to eukaryotic cells. Peptide transport systems often consist of a substrate-binding protein associated with a membrane-spanning permease. The environmental bacterial pathogen Listeria monocytogenes uses the substrate-binding protein CtaP not only for cysteine transport but also for resistance to acid, maintenance of membrane integrity, and bacterial adherence to host cells. In this study, we demonstrate complementary yet distinct functional roles for two membrane permeases, CtpP1 and CtpP2, that are encoded by genes linked to ctaP and that contribute to bacterial growth, invasion, and pathogenicity.

Asunto(s)

Listeria monocytogenes , Animales , Ratones , Listeria monocytogenes/genética , Cisteína/metabolismo , Virulencia , Proteínas de Transporte de Membrana/genética , Proteínas de Transporte de Membrana/metabolismo , Factores de Virulencia/genética , Proteínas Bacterianas/metabolismo , Modelos Animales de Enfermedad , Regulación Bacteriana de la Expresión Génica , Mamíferos

RESUMEN

A number of bacterial pathogens are capable of detecting the presence of other bacteria located within their surrounding niche through a process of bacterial signaling and cell-to-cell communication commonly referred to as quorum sensing (QS). QS systems are commonly now described in the context of collective behaviors exhibited by groups of bacteria coordinating diverse arrays of physiological functions to enhance survival of the community. However, QS systems have also been implicated in a variety of processes distinct from the measure of bacterial cell density. This review will highlight noncanonical adaptations of canonical QS systems that have evolved to enable bacteria to detect nonself individuals within a population or to detect occupation of confined spaces.

Asunto(s)

Bacterias/metabolismo , Fenómenos Fisiológicos Bacterianos , Percepción de Quorum/fisiología , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Comunicación Celular/fisiología , Recuento de Células , Enterococcus/genética , Enterococcus/metabolismo , Regulación Bacteriana de la Expresión Génica , Interacciones Huésped-Parásitos/fisiología , Listeria/genética , Listeria/metabolismo , Transducción de Señal/fisiología , Staphylococcus/genética , Staphylococcus/metabolismo , Vacuolas/microbiología

RESUMEN

Anesthetics are known to modulate host immune responses, but separating the variables of surgery from anesthesia when analyzing hospital acquired infections is often difficult. Here, the bacterial pathogen Listeria monocytogenes (Lm) was used to assess the impact of the common anesthetic propofol on host susceptibility to infection. Brief sedation of mice with physiologically relevant concentrations of propofol increased bacterial burdens in target organs by more than 10,000-fold relative to infected control animals. The adverse effects of propofol sedation on immune clearance of Lm persisted after recovery from sedation, as animals given the drug remained susceptible to infection for days following anesthesia. In contrast to propofol, sedation with alternative anesthetics such as ketamine/xylazine or pentobarbital did not increase susceptibility to systemic Lm infection. Propofol altered systemic cytokine and chemokine expression during infection, and prevented effective bacterial clearance by inhibiting the recruitment and/or activity of immune effector cells at sites of infection. Propofol exposure induced a marked reduction in marginal zone macrophages in the spleens of Lm infected mice, resulting in bacterial dissemination into deep tissue. Propofol also significantly increased mouse kidney abscess formation following infection with the common nosocomial pathogen Staphylococcus aureus. Taken together, these data indicate that even brief exposure to propofol severely compromises host resistance to microbial infection for days after recovery from sedation.

Asunto(s)

Infecciones Bacterianas/inducido químicamente , Infecciones Bacterianas/inmunología , Susceptibilidad a Enfermedades/inducido químicamente , Hipnóticos y Sedantes/efectos adversos , Macrófagos/efectos de los fármacos , Propofol/efectos adversos , Linfocitos T/efectos de los fármacos , Animales , Animales no Consanguíneos , Infecciones Bacterianas/patología , Recuento de Células , Células Cultivadas , Infección Hospitalaria/inducido químicamente , Infección Hospitalaria/inmunología , Infección Hospitalaria/patología , Susceptibilidad a Enfermedades/inmunología , Regulación hacia Abajo/efectos de los fármacos , Regulación hacia Abajo/inmunología , Femenino , Listeria monocytogenes/efectos de los fármacos , Listeria monocytogenes/crecimiento & desarrollo , Listeriosis/inducido químicamente , Listeriosis/inmunología , Listeriosis/patología , Macrófagos/inmunología , Macrófagos/patología , Ratones , Linfocitos T/inmunología , Linfocitos T/patología

RESUMEN

Synthesis of the cyclic nucleotide c-di-AMP has been reported for a number of bacteria, but the physiological role of this apparently essential second messenger has remained unclear. In this issue of Cell Host & Microbe, Whiteley et al. (2015) elucidate the pathways linking c-di-AMP with the appropriate regulation of bacterial metabolism.

Asunto(s)

Fosfatos de Dinucleósidos/metabolismo , Guanosina Pentafosfato/metabolismo , Listeria monocytogenes/crecimiento & desarrollo , Animales , Femenino

RESUMEN

Listeria monocytogenes is a Gram-positive facultative intracellular bacterial pathogen that invades mammalian cells and escapes from membrane-bound vacuoles to replicate within the host cell cytosol. Gene products required for intracellular bacterial growth and bacterial spread to adjacent cells are regulated by a transcriptional activator known as PrfA. PrfA becomes activated following L. monocytogenes entry into host cells, however the signal that stimulates PrfA activation has not yet been defined. Here we provide evidence for L. monocytogenes secretion of a small peptide pheromone, pPplA, which enhances the escape of L. monocytogenes from host cell vacuoles and may facilitate PrfA activation. The pPplA pheromone is generated via the proteolytic processing of the PplA lipoprotein secretion signal peptide. While the PplA lipoprotein is dispensable for pathogenesis, bacteria lacking the pPplA pheromone are significantly attenuated for virulence in mice and have a reduced efficiency of bacterial escape from the vacuoles of nonprofessional phagocytic cells. Mutational activation of PrfA restores virulence and eliminates the need for pPplA-dependent signaling. Experimental evidence suggests that the pPplA peptide may help signal to L. monocytogenes its presence within the confines of the host cell vacuole, stimulating the expression of gene products that contribute to vacuole escape and facilitating PrfA activation to promote bacterial growth within the cytosol.

Asunto(s)

Evasión Inmune , Lipoproteínas/inmunología , Listeria monocytogenes/inmunología , Péptidos/inmunología , Feromonas/inmunología , Vacuolas/inmunología , Animales , Proteínas Bacterianas/genética , Proteínas Bacterianas/inmunología , Lipoproteínas/genética , Listeria monocytogenes/genética , Ratones , Factores de Terminación de Péptidos/genética , Factores de Terminación de Péptidos/inmunología , Péptidos/genética , Feromonas/genética , Vacuolas/microbiología

RESUMEN

Environmental pathogens - organisms that survive in the outside environment but maintain the capacity to cause disease in mammals - navigate the challenges of life in habitats that range from water and soil to the cytosol of host cells. The bacterium Listeria monocytogenes has served for decades as a model organism for studies of host-pathogen interactions and for fundamental paradigms of cell biology. This ubiquitous saprophyte has recently become a model for understanding how an environmental bacterium switches to life within human cells. This review describes how L. monocytogenes balances life in disparate environments with the help of a critical virulence regulator known as PrfA. Understanding L. monocytogenes survival strategies is important for gaining insight into how environmental microbes become pathogens.

Asunto(s)

Proteínas Bacterianas/metabolismo , Microbiología Ambiental , Regulación Bacteriana de la Expresión Génica/genética , Listeria monocytogenes/fisiología , Listeriosis/microbiología , Factores de Terminación de Péptidos/metabolismo , Proteínas Bacterianas/genética , Interacciones Huésped-Patógeno , Humanos , Listeria monocytogenes/genética , Listeria monocytogenes/patogenicidad , Modelos Moleculares , Factores de Terminación de Péptidos/genética , Factores de Virulencia

RESUMEN

The environmental bacterium Listeria monocytogenes survives and replicates in a variety of diverse ecological niches that range from the soil to the cytosol of infected mammalian cells. The ability of L. monocytogenes to replicate within an infected host requires the expression of a number of secreted bacterial gene products whose expression is regulated by the transcriptional activator PrfA. PrfA becomes activated following bacterial entry into host cells; however, the mechanism by which this activation occurs remains unknown. Here we describe a novel C-terminal mutation that results in the high-level constitutive activation of PrfA and yet, in contrast with other described prfA* activation mutations, only modestly increases PrfA DNA binding affinity. L. monocytogenes strains containing the prfA P219S mutation exhibited high levels of PrfA-dependent virulence gene expression, were hyperinvasive in tissue culture models of infection, were fully motile and were hypervirulent in mice. In contrast with PrfA G145S and other mutationally activated PrfA proteins, the PrfA P219S protein readily formed homodimers and did not exhibit a dramatic increase in its DNA-binding affinity for target promoters. Interestingly, the prfA P219S mutation is located adjacent to the prfA K220 residue that has been previously reported to contribute to PrfA DNA binding activity. prfA P219S therefore appears to constitutively activate PrfA via a novel mechanism which minimally affects PrfA DNA binding in vitro.

Asunto(s)

Proteínas Bacterianas/metabolismo , Listeria monocytogenes/genética , Factores de Terminación de Péptidos/metabolismo , Factores de Virulencia/metabolismo , Animales , Proteínas Bacterianas/genética , Células Cultivadas , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Femenino , Regulación Bacteriana de la Expresión Génica , Listeria monocytogenes/patogenicidad , Ratones , Mutación , Factores de Terminación de Péptidos/genética , Estructura Terciaria de Proteína , Factores de Virulencia/genética

RESUMEN

Listeria monocytogenes is a food-borne intracellular bacterial pathogen capable of causing serious human disease. L. monocytogenes survival within mammalian cells depends upon the synthesis of a number of secreted virulence factors whose expression is regulated by the transcriptional activator PrfA. PrfA becomes activated following bacterial entry into host cells where it induces the expression of gene products required for bacterial spread to adjacent cells. Activation of PrfA appears to occur via the binding of a small molecule cofactor whose identity remains unknown. Electrostatic modeling of the predicted PrfA cofactor binding pocket revealed a highly positively charged region with two lysine residues, K64 and K122, located at the edge of the pocket and another (K130) located deep within the interior. Mutational analysis of these residues indicated that K64 and K122 contribute to intracellular activation of PrfA, whereas a K130 substitution abolished protein activity. The requirement of K64 and K122 for intracellular PrfA activation could be bypassed via the introduction of the prfA G145S mutation that constitutively activates PrfA in the absence of cofactor binding. Our data indicate that the positive charge of the PrfA binding pocket contributes to intracellular activation of PrfA, presumably by facilitating binding of an anionic cofactor.

Asunto(s)

Proteínas Bacterianas/metabolismo , Listeria monocytogenes/metabolismo , Listeriosis/microbiología , Factores de Terminación de Péptidos/metabolismo , Sustitución de Aminoácidos , Animales , Proteínas Bacterianas/química , Proteínas Bacterianas/genética , Sitios de Unión/genética , Western Blotting , Humanos , Listeria monocytogenes/genética , Listeria monocytogenes/patogenicidad , Listeriosis/patología , Lisina/química , Lisina/genética , Lisina/metabolismo , Ratones , Modelos Moleculares , Mutación , Factores de Terminación de Péptidos/química , Factores de Terminación de Péptidos/genética , Unión Proteica , Conformación Proteica , Estructura Terciaria de Proteína , Propiedades de Superficie , Virulencia/genética

RESUMEN

As an organism that has evolved to live in environments ranging from soil to the cytosol of mammalian cells, Listeria monocytogenes must regulate the secretion and activity of protein products that promote survival within these habitats. The post-translocation chaperone PrsA2 has been adapted to assist in the folding and activity of L. monocytogenes secreted proteins required for bacterial replication within host cells. Here we present the first structure/function investigation of the contributions of PrsA2 to protein secretion and activity as well as to bacterial virulence. Domain swap experiments with the closely related L. monocytogenes PrsA1 protein combined with targeted mutagenesis indicate distinct functional roles for the PrsA2 peptidyl-prolyl isomerase (PPIase) and the N- and C-terminal domains in pathogenesis. In contrast to other PrsA-like proteins described thus far in the literature, an absolute in vivo requirement for PrsA2 PPIase activity is evident in mouse infection models. This work illustrates the diversity of function associated with L. monocytogenes PrsA2 that serves to promote bacterial life within the infected host.

Asunto(s)

Listeria monocytogenes/enzimología , Listeria monocytogenes/patogenicidad , Listeriosis/microbiología , Isomerasa de Peptidilprolil/metabolismo , Secuencia de Aminoácidos , Animales , Femenino , Regulación Bacteriana de la Expresión Génica , Humanos , Listeria monocytogenes/química , Listeria monocytogenes/genética , Ratones , Conformación Molecular , Datos de Secuencia Molecular , Isomerasa de Peptidilprolil/química , Isomerasa de Peptidilprolil/genética , Estructura Terciaria de Proteína , Alineación de Secuencia , Virulencia

RESUMEN

Listeria monocytogenes secretes two chitinases and one chitin binding protein. Mutants lacking chiA, chiB, or lmo2467 exhibited normal growth in cultured cells but were defective for growth in the livers and spleens of mice. Mammals lack chitin; thus, L. monocytogenes may have adapted chitinases to recognize alternative substrates to enhance pathogenesis.

Asunto(s)

Quitinasas/fisiología , Listeria monocytogenes/patogenicidad , Animales , Células CACO-2/microbiología , Quitina/metabolismo , Quitinasas/genética , Quitinasas/metabolismo , Genes Bacterianos/genética , Humanos , Listeria monocytogenes/enzimología , Listeria monocytogenes/genética , Listeriosis/microbiología , Hígado/microbiología , Ratones , Bazo/microbiología

RESUMEN

Riboswitches are RNA structures traditionally viewed as acting in cis to regulate downstream gene expression in bacteria. In a recent issue of Cell, Loh and colleagues report on the ability of a riboswitch to act in trans to modulate the expression of a critical bacterial virulence regulator.

RESUMEN

The bacterial pathogen Listeria monocytogenes survives under a myriad of conditions in the outside environment and within the human host where infections can result in severe disease. Bacterial life within the host requires the expression of genes with roles in nutrient acquisition as well as the biosynthesis of bacterial products required to support intracellular growth. A gene product identified as the substrate-binding component of a novel oligopeptide transport system (encoded by lmo0135) was recently shown to be required for L. monocytogenes virulence. Here we demonstrate that lmo0135 encodes a multifunctional protein that is associated with cysteine transport, acid resistance, bacterial membrane integrity and adherence to host cells. The lmo0135 gene product (designated CtaP, for cysteine transport associated protein) was required for bacterial growth in the presence of low concentrations of cysteine in vitro, but was not required for bacterial replication within the host cytosol. Loss of CtaP increased membrane permeability and acid sensitivity, and reduced bacterial adherence to host cells. ctaP deletion mutants were severely attenuated following intragastric and intravenous inoculation of mice. Taken together, the data presented indicate that CtaP contributes to multiple facets of L. monocytogenes physiology, growth and survival both inside and outside of animal cells.

Asunto(s)

Proteínas Bacterianas/fisiología , Cisteína/metabolismo , Listeria monocytogenes/patogenicidad , Proteínas de Transporte de Membrana/fisiología , Factores de Virulencia/fisiología , Ácidos/toxicidad , Estructuras Animales/microbiología , Animales , Adhesión Bacteriana , Proteínas Bacterianas/genética , Permeabilidad de la Membrana Celular/fisiología , Recuento de Colonia Microbiana , Femenino , Eliminación de Gen , Listeriosis/microbiología , Ratones , Viabilidad Microbiana , Modelos Biológicos , Estrés Fisiológico , Factores de Virulencia/genética

RESUMEN

Extracellular polysaccharides of many bacteria are synthesized by the Wzy polymerase-dependent mechanism, where long-chain polymers are assembled from undecaprenyl-phosphate-linked repeat units on the outer face of the cytoplasmic membrane. In gram-positive bacteria, Wzy-dependent capsules remain largely cell associated via membrane and peptidoglycan linkages. Like many Wzy-dependent capsules, the Streptococcus pneumoniae serotype 2 capsule is branched. In this study, we found that deletions of cps2K, cps2J, or cps2H, which encode a UDP-glucose dehydrogenase necessary for side chain synthesis, the putative Wzx transporter (flippase), and the putative Wzy polymerase, respectively, were obtained only in the presence of suppressor mutations. Most of the suppressor mutations were in cps2E, which encodes the initiating glycosyltransferase for capsule synthesis. The cps2K mutants containing the suppressor mutations produced low levels of high-molecular-weight polymer that was detected only in membrane fractions. cps2K-repaired mutants exhibited only modest increases in capsule production due to the effect of the secondary mutation, but capsule was detectable in both membrane and cell wall fractions. Lethality of the cps2K, cps2J, and cps2H mutations was likely due to sequestration of undecaprenyl-phosphate in the capsule pathway and either preclusion of its turnover for utilization in essential pathways or destabilization of the membrane due to an accumulation of lipid-linked intermediates. The results demonstrate that proper polymer assembly requires not only a functional transporter and polymerase but also complete repeat units. A central role for the initiating glycosyltransferase in controlling capsule synthesis is also suggested.

Asunto(s)

Cápsulas Bacterianas/metabolismo , Pared Celular/metabolismo , Genes Esenciales , Mutación , Polisacáridos Bacterianos/metabolismo , Streptococcus pneumoniae/fisiología , Cápsulas Bacterianas/genética , Proteínas Bacterianas/genética , Fraccionamiento Celular , Membrana Celular/química , Eliminación de Gen , Glicosiltransferasas/genética , Proteínas de Transporte de Membrana/genética , Viabilidad Microbiana , Modelos Moleculares , Fosfatos de Poliisoprenilo/metabolismo , Polisacáridos Bacterianos/genética , Streptococcus pneumoniae/genética , Streptococcus pneumoniae/crecimiento & desarrollo , Supresión Genética , Uridina Difosfato Glucosa Deshidrogenasa/genética