Biomedical subjects

Pascale Cossart

Publications and source records attributed to Pascale Cossart.

63 records · Page 4Linked to original sources

PubMed2002

An RNA thermosensor controls expression of virulence genes in Listeria monocytogenes.

In Listeria monocytogenes, virulence genes are maximally expressed at 37 degrees C, almost silent at 30 degrees C and controlled by PrfA, a transcriptional activator whose expression is thermoregulated. Here, we show that the untranslated mRNA (UTR) preceding prfA, forms a secondary structure, which masks the ribosome binding region. Mutations predicted to destabilize this structure led to virulence gene expression and invasion of mammalian cells at 30 degrees C. Chemical probing, native gel electrophoresis, in vitro translation, and "compensatory" and "increased stability" mutations demonstrated that the UTR switches between a structure active at high temperatures, and another inactive at low temperatures. Strikingly, when the DNA corresponding to the UTR was fused to gfp in E. coli, bacteria became fluorescent at 37 degrees C, but not at 30 degrees C. This mechanism of posttranscriptional thermoregulation may have important applications.

Bacterial Proteins↗

PubMed2002

InlB, a surface protein of Listeria monocytogenes that behaves as an invasin and a growth factor.

Molecules from some pathogenic bacteria mimic natural host cell ligands and trigger engulfment of the bacterium after specifically interacting with cell-surface receptors. The leucine-rich repeat (LRR)-containing protein InlB of Listeria monocytogenes is one such molecule. It triggers bacterial entry by interacting with the hepatocyte growth factor receptor (HGF-R or Met) and two other cellular components: gC1q-R and proteoglycans. Recent studies point to significant similarities between the molecular mechanisms underlying InlB-mediated entry into cells and classic phagocytosis. In addition, InlB, in common with HGF, activates signaling cascades that are not involved in bacterial entry. Therefore, studies of InlB may help us to analyze the previously noticed similarities between growth factor receptor activation and phagocytosis.

Actins↗

PubMed2002

Listeriolysin O: a genuine cytolysin optimized for an intracellular parasite.

Cholesterol-dependent cytolysins (CDCs)* are produced by a large number of pathogenic gram-positive bacteria. A member of this family, listeriolysin O (LLO), is produced by the intracellular pathogen Listeria monocytogenes. A unique feature of LLO is its low optimal pH activity (approximately 6) which permits escape of the bacterium from the phagosome into the host cell cytosol without damaging the plasma membrane of the infected cell. In a recent study (Glomski et al., 2002, this issue), Portnoy's group has addressed the molecular mechanism underlying the pH sensitivity of LLO. Unexpectedly, a single amino acid substitution in LLO L461T results in a molecule more active at neutral pH and promoting premature permeabilization of the infected cells, leading to attenuated virulence. This finding highlights how subtle changes in proteins can be exploited by bacterial pathogens to establish and maintain the integrity of their specific niches.

Animals↗

PubMed2002

Surface proteins and the pathogenic potential of Listeria monocytogenes.

On the basis of the recently determined genome sequence of Listeria monocytogenes, we performed a global analysis of the surface-protein-encoding genes. Only proteins displaying a signal peptide were taken into account. Forty-one genes encoding LPXTG proteins, including the previously known internalin gene family, were detected. Several genes encoding proteins that, like InlB and Ami, possess GW modules that attach them to lipoteichoic acids were also identified. Additionally, the completed genome sequence revealed genes encoding proteins potentially anchored in the cell membrane by a hydrophobic tail as well as genes encoding P60-like proteins and lipoproteins. We describe these families and discuss their putative implications for host-pathogen interactions.

Amino Acid Motifs↗

PubMed2002

Genetically-modified-animal models for human infections: the Listeria paradigm.

Several human pathogens exhibit a restricted host-tropism, relying on the species-specific interaction of microbial ligand(s) with host receptor(s). This specificity accounts for some of the difficulties in modeling human infections in animals. The discovery of L. monocytogenes host-specificity and elucidation of the underlying mechanism has led to the generation of transgenic mice expressing one of its human receptors, E-cadherin. This model is presented here as a paradigm of a genetically-modified-animal model for studying a human infectious disease.

Animals↗

PubMed2002

Inactivation of the srtA gene in Listeria monocytogenes inhibits anchoring of surface proteins and affects virulence.

During infection of their hosts, Gram-positive bacteria express surface proteins that serve multiple biological functions. Surface proteins harbouring a C-terminal sorting signal with an LPXTG motif are covalently linked to the cell wall peptidoglycan by a transamidase named sortase. Two genes encoding putative sortases, termed srtA and srtB, were identified in the genome of the intracellular pathogenic bacterium Listeria monocytogenes. Inactivation of srtA abolishes anchoring of the invasion protein InlA to the bacterial surface. It also prevents the proper sorting of several other peptidoglycan-associated LPXTG proteins. Three were identified by a mass spectrometry approach. The DeltasrtA mutant strain is defective in entering epithelial cells, similar to a DeltainlA mutant. In contrast to a DeltainlA mutant, the DeltasrtA mutant is impaired for colonization of the liver and spleen after oral inoculation in mice. Thus, L. monocytogenes srtA is required for the cell wall anchoring of InlA and, presumably, for the anchoring of other LPXTG-containing proteins that are involved in listerial infections.

Amino Acid Sequence↗

PubMed2002

Listeria monocytogenes bile salt hydrolase is a PrfA-regulated virulence factor involved in the intestinal and hepatic phases of listeriosis.

Listeria monocytogenes is a bacterial pathogen causing severe food-borne infections in humans and animals. It can sense and adapt to a variety of harsh microenvironments outside as well as inside the host. Once ingested by a mammalian host, the bacterial pathogen reaches the intestinal lumen, where it encounters bile salts which, in addition to their role in digestion, have antimicrobial activity. Comparison of the L. monocytogenes and Listeria innocua genomes has revealed the presence of an L. monocytogenes-specific putative gene encoding a bile salt hydrolase (BSH). Here, we show that the bsh gene encodes a functional intracellular enzyme in all pathogenic Listeria species. The bsh gene is positively regulated by PrfA, the transcriptional activator of known L. monocytogenes virulence genes. Moreover, BSH activity increases at low oxygen concentration. Deletion of bsh results in decreased resistance to bile in vitro, reduced bacterial faecal carriage after oral infection of the guinea-pigs, reduced virulence and liver colonization after intravenous inoculation of mice. Taken together, these results demonstrate that BSH is a novel PrfA-regulated L. monocytogenes virulence factor involved in the intestinal and hepatic phases of listeriosis.

Amidohydrolases↗

PubMed2002

Distinct protein patterns associated with Listeria monocytogenes InlA- or InlB-phagosomes.

Internalization of Listeria monocytogenes into non-phagocytic cells is mediated by the interactions between the two bacterial invasion proteins InlA (internalin) and InlB and their cellular surface receptors E-cadherin and c-Met. To get an insight into all the cellular components necessary for uptake and early intracellular life, we undertook a global proteomic characterization of the early listerial phagosome in the human epithelial cell line LoVo. First, we proceeded to an immunocytochemical characterization of intracellular marker recruitment to phagosomes containing latex beads coated with InlA or InlB. E-cadherin and c-Met were, as expected, rapidly recruited to the phagosomal formation site. Phagosomes subsequently acquired the early endosomal antigen 1 (EEA1) and the lysosomal-associated membrane protein 1 (LAMP1), while presenting a more delayed enrichment of the lysosomal hydrolase cathepsin D. Early phagosomes devoid of lysosomal, endoplasmic reticulum and Golgi enzymatic activities could then be isolated by subcellular fractionation of LoVo cells. Two-dimensional gel electrophoresis (2DPAGE) revealed differences between the protein profiles of InlA- or InlB-phagosomes and those of early/late endosomes or lysosomes of naive LoVo cells. One major protein specifically recruited on the InlB-phagosomes was identified by mass spectrometry as MSF, a previously reported member of the septin family of GTPases. MSF forms filaments that co-localize with the actin cytoskeleton in resting cells and it is recruited to the entry site of InlB-coated beads. These results suggest that MSF is a putative effector of the InlB-mediated internalization of L. monocytogenes into host cells.

Actins↗

PubMed2002

Molecular and cellular basis of the infection by Listeria monocytogenes: an overview.

This review rather than covering the whole field, intends to highlight the particularly interesting properties of some proteins involved in the infection by Listeria monocytogenes and the general interest in some of the approaches used to analyze the molecular and cellular basis of the infection. After an introduction to the bacterium and to the disease, a description of the infection at the cellular level will be given. The specific features of the pore-forming toxin listeriolysin O, as a protein particularly well adapted to the intracellular lifestyle of L. monocytogenes will be discussed. By describing in detail how the bacterium moves inside cells, particular attention will be given to show how addressing this issue has provided key answers and instrumental tools to cell biologists studying actin-based motility. The analysis of the entry process and in particular the studies derived from the specificity of internalin for its receptor will demonstrate how an apparently reductionist approach can help generating relevant animal models, identification of virulence factors and demonstration of their role in vivo. The virulence gene cluster and its regulation by PrfA will be presented and discussed in the framework of the recently determined genome sequence.

Actins↗