Catherine Gouyette

Virulence strategies for infecting phagocytes deduced from the in vivo transcriptional program of Legionella pneumophila

Adaptation to the host environment and exploitation of host cell functions are critical to the success of intracellular pathogens. Here, insight to these virulence mechanisms was obtained for the first time from the transcriptional program of the human pathogen Legionella pneumophila during infection of its natural host, Acanthamoeba castellanii. The biphasic life cycle of L. pneumophila was reflected by a major shift in gene expression from replicative to transmissive phase, concerning nearly half of the genes predicted in the genome. However, three different L. pneumophila strains showed similar in vivo gene expression patterns, indicating that common regulatory mechanisms govern the Legionella life cycle, despite the plasticity of its genome. During the replicative phase, in addition to components of aerobic metabolism and amino acid catabolism, the Entner‐Doudoroff pathway, a NADPH producing mechanism used for sugar and/or gluconate assimilation, was expressed, suggesting for the first time that intracellular L. pneumophila may also scavenge host carbohydrates as nutrients and not only proteins. Identification of genes only upregulated in vivo but not in vitro, may explain higher virulence of in vivo grown L. pneumophila. Late in the life cycle, L. pneumophila upregulates genes predicted to promote transmission and manipulation of a new host cell, therewith priming it for the next attack. These including substrates of the Dot/Icm secretion system, other factors associated previously with invasion and virulence, the motility and the type IV pilus machineries, and > 90 proteins not characterized so far. Analysis of a fliA (σ28) deletion mutant identified genes coregulated with the flagellar regulon, including GGDEF/EAL regulators and factors that promote host cell entry and survival.

Stress by Heat Shock Induces Massive Down Regulation of Genes and Allows Differential Allelic Expression of the Gal/GalNAc Lectin in Entamoeba histolytica

ABSTRACT Gene expression analysis by microarray assay revealed that when exposed to stress, Entamoeba histolytica exhibits a specific heat shock response, together with a dramatic overall reduction in gene transcription as well as differential allelic expression of key genes participating in virulence, such as the galactose/N-acetylgalactosamine (Gal/GalNAc) lectin.

Genomic polymorphism in the population of Candida glabrata: Gene copy-number variation and chromosomal translocations

The genomic sequence of the type strain of the opportunist human pathogen Candida glabrata (CBS138, ATCC 2001) is available since 2004. This allows the analysis of genomic structure of other strains by comparative genomic hybridization. We present here the molecular analysis of a collection of 183 C. glabrata strains isolated from patients hospitalized in France and around the world. We show that the mechanisms of microevolution within this asexual species include rare reciprocal chromosomal translocations and recombination within tandem arrays of repeated genes, and that these account for the frequent size heterogeneity between chromosomes across strains. Gene tandems often encode cell wall proteins suggesting a possible role in adaptation to the environment.

The interaction of manganese ions with DNA

We present the structure of the duplex formed by a fragment of auto-complementary DNA with the sequence d(CGTTAATTAACG). The structure was determined by X-ray crystallography. Up to date it is the first structure presenting the interaction between a DNA oligonucleotide and manganese ions. The presence of Mn2+ creates bonds among the N7 atom of guanines and phosphates. These bonds stabilize and determine the crystallographic network in a P3(2) space group, unusual in oligonucleotide crystals. The crystal structure observed is compared with those found in the presence of Mg2+, Ca2+ and Ni2+, which show different kinds of interactions. The double helices show end-to-end interactions, in a manner that the terminal guanines interact with the minor groove of the neighboring duplex, while the terminal cytosines are disordered. We have chosen this sequence since it contains a TTAA repeat. Such repeats are very rare in all genomes. We suggest that this sequence may be very susceptible to the formation of closely spaced thymine dimers.

6- and 1- substituted mannosyl phosphotriesters as lipophilic macrophage-targeted carriers of antiviral nucleosides

Abstract Lipophilic mannose-6 phosphate and (1-α mannosyl)ethyl phosphate derivatives of the antiviral nucleosides 3′-azido thymidine (AZT), 2′,3′-dideoxy thymidine (ddT) and 2′-deoxy 5-fluoro uridine (dU F ) were prepared from D-mannose as membrane soluble prodrugs directed towards cells carrying mannosyl receptors.

Synthesis of oligonucleotide–peptide conjugates using hydrazone chemical ligation

An oligonucleotide was functionalized on the solid-phase by a tartaramide moiety, which could be converted efficiently in solution into a glyoxylyl group following a mild periodic oxidation. The glyoxylyl-oligonucleotide was found to be very stable upon storage and was successfully engaged in hydrazone ligation with an α-hydrazino acetyl peptide.

Lactobacillus gigeriorum sp. nov., isolated from chicken crop.

In the early 1980s, a facultatively anaerobic, non-motile, short rod, designated 202(T), was isolated from a chicken crop and identified as a homofermentative lactic acid bacterium. Phylogenetic analysis based on the 16S rRNA gene sequence revealed that the strain was affiliated with the genus Lactobacillus, clustering within the Lactobacillus acidophilus-delbrueckii group. In this analysis, strain 202(T) appeared to be most closely related to the type strains of Lactobacillus intestinalis and Lactobacillus amylolyticus, with gene sequence similarities of 96.1 and 96.2 %, respectively. Strain 202(T) was found to differ from these two species, however, when investigated by multilocus sequence analysis, and it also differed in terms of some of its metabolic properties. On the basis of these observations, strain 202(T) is considered to represent a novel species in the genus Lactobacillus, for which the name Lactobacillus gigeriorum sp. nov. is proposed; the type strain is 202(T) ( = CRBIP 24.85(T) = DSM 23908(T)).

Spectroscopic evidence for an intramolecular RNA triple helix

A 29‐base RNA oligomer has been chemically synthesized and shown to form an intramolecular triple helix in solution at acidic pH. Assignment of the majority of the exchangeable proton NMR resonances demonstrated the Watson—Crick and Hoogsteen base pairings consistent with folding to form pyrimidine—purine—pyrimidine base triplets. FTIR spectroscopy provided independent evidence of base triplet formation, and indicated a predominately C3′‐endo sugar pucker. UV absorption as a function of temperature suggested monophasic melting behaviour, which was confirmed by NMR of the imino protons.

Synthese de C-nucleosides—IX : Purines substituees en position 8 par le desoxy-2 ribose

Abstract Benzyl 2-deoxy-3,5-di-O-p-toluyl- d - erythro -pentofuranosyl thioformimidate 3 , prepared from nitrile 1 , reacts with α-aminocyanoacetic acid derivatives to yield C-imidazole nucleosides which are further cyclized into purines. The 6-mercapto purine 2 is obtained in two different ways.

Structure of the DNA Coiled Coil Formed by d(CGATATATATAT)

AT-rich sequences of DNA are highly polymorphic, as we have reviewed elsewhere and are abundant in noncoding regions of the genome. As part of our studies on these sequences, we have determined the molecular structure of the coiled coil obtained from duplex d(CGATATATATAT) by single-crystal X-ray diffraction. We recently described the structure of the coiled coil formed by duplex d(ATATATATATAT). Due to the presence of sticky ends it forms a continuous double helix. However diffraction data could only be obtained with a limited resolution of 5 1; this is probably due to a combination of factors: high solvent content, screw disorder, and possible multiple alignments. In order to eliminate the latter factor, we crystallized d(CGATATATATAT), which can only form stacked infinite continuous duplexes in a single way, as shown in Figure 1. The crystals were practically isomorphous to those previously described. We determined the 3.1 1 X-ray structure and found that this dodecamer forms a continuous coiled coil with Hoogsteen base pairs between A and T.