InhA1-Mediated Cleavage of the Metalloprotease NprA Allows to Escape From Macrophages.

is a Gram-positive spore-forming bacterium causing food poisoning and serious opportunistic infections. These infections are characterized by bacterial accumulation in the host despite the induction of inflammation. To circumvent inflammation, bacteria must resist the bactericidal activity of professional phagocytes, which constitute a first line of host defense against pathogens.

The Bacterial Mfd Protein Prevents DNA Damage Induced by the Host Nitrogen Immune Response in a NER-Independent but RecBC-Dependent Pathway.

Production of reactive nitrogen species is an important component of the host immune defence against bacteria. Here, we show that the bacterial protein Mfd (Mutation frequency decline), a highly conserved and ubiquitous bacterial protein involved in DNA repair, confers bacterial resistance to the eukaryotic nitrogen response produced by macrophage cells and during mice infection. In addition, we show that RecBC is also necessary to survive this stress.

The bacterial DNA repair protein Mfd confers resistance to the host nitrogen immune response.

Production of reactive nitrogen species (NO) is a key step in the immune response following infections. NO induces lesions to bacterial DNA, thus limiting bacterial growth within hosts. Using two pathogenic bacteria, Bacillus cereus and Shigella flexneri, we show that the DNA-repair protein Mfd (Mutation-Frequency-Decline) is required for bacterial resistance to the host-NO-response.

Iron regulates Bacillus thuringiensis haemolysin hlyII gene expression during insect infection.

Bacillus thuringiensis (Bt) is a spore-forming entomopathogen broadly used in agriculture crop. The haemolysin HlyII is an important Bt virulence factor responsible for insect death. In this work, we focused on the regulation of the hlyII gene throughout the bacterial growth in vitro and in vivo during insect infection.

Glucose 6P binds and activates HlyIIR to repress Bacillus cereus haemolysin hlyII gene expression.

Bacillus cereus is a Gram-positive spore-forming bacterium causing food poisoning and serious opportunistic infections. These infections are characterized by bacterial accumulation despite the recruitment of phagocytic cells. We have previously shown that B.

Haemolysin II is a Bacillus cereus virulence factor that induces apoptosis of macrophages.

Bacillus cereus is a Gram-positive spore-forming bacterium causing food poisoning and serious opportunistic infections. These infections are characterized by bacterial accumulation despite the recruitment of phagocytic cells. The precise mechanisms and the bacterial factors allowing B.

CwpFM (EntFM) is a Bacillus cereus potential cell wall peptidase implicated in adhesion, biofilm formation, and virulence.

Bacillus cereus EntFM displays an NlpC/P60 domain, characteristic of cell wall peptidases. The protein is involved in bacterial shape, motility, adhesion to epithelial cells, biofilm formation, vacuolization of macrophages, and virulence. These data provide new information on this, so far, poorly studied toxin and suggest that this protein is a cell wall peptidase, which we propose to rename CwpFM.

The InhA metalloproteases of Bacillus cereus contribute concomitantly to virulence.

The virulence of Bacillus cereus requires that bacteria have the capacity to colonize their host, degrade specific tissues, and circumvent the host immune system. To study this aspect of pathogenesis, we focused on three metalloproteases, InhA1, InhA2, and InhA3, which share more than 66% identity. The expression of these metalloprotease genes was assessed by transcriptional fusions with a lacZ reporter gene.

Characterization of two Bacillus thuringiensis genes identified by in vivo screening of virulence factors.

Bacillus thuringiensis vegetative cells are known to be highly pathogenic when injected into the hemocoel of susceptible insect larvae. This pathogenicity is due to the capacity of B. thuringiensis to cause septicemia in the host.

The [URE3] phenotype: evidence for a soluble prion in yeast.

The aggregation of the two yeast proteins Sup35p and Ure2p is widely accepted as a model for explaining the prion propagation of the phenotypes [PSI+] and [URE3], respectively. Here, we demonstrate that the propagation of [URE3] cannot simply be the consequence of generating large aggregates of Ure2p, because such aggregation can be found in some conditions that are not related to the prion state of Ure2p. A comparison of [PSI+] and [URE3] aggregation demonstrates differences between these two prion mechanisms.

The yeast prion [URE3] can be greatly induced by a functional mutated URE2 allele.

The non-Mendelian element [URE3] of yeast is considered to be a prion form of the Ure2 protein. The [URE3] phenotype occurs at a frequency of 10(-5) in haploid yeast strains, is reversible, and its frequency is increased by overexpressing the URE2 gene. We created a new mutant of the Ure2 protein, called H2p, which results in a 1000-fold increase in the rate of [URE3] occurrence.

Characterization of the interaction domains of Ure2p, a prion-like protein of yeast.

In the yeast Saccharomyces cerevisiae, the non-Mendelian inherited genetic element [URE3] behaves as a prion. A hypothesis has been put forward which states that [URE3] arises spontaneously from its cellular isoform Ure2p (the product of the URE2 gene), and propagates through interactions of the N-terminal domain of the protein, thus leading to its aggregation and loss of function. In the present study, various N- and C-terminal deletion mutants of Ure2p were constructed and their cross-interactions were tested in vitro and in vivo using affinity binding and a two-hybrid analysis.

Enhanced expression of the yeast Ure2 protein in Escherichia coli: the effect of synonymous codon substitutions at a selected place in the gene.

The expression of the yeast Ure2 protein and its two N- and C-terminal HA-(YPYPVDYA) epitope and His-tag fusions has been enhanced in E. coli by selected silent mutagenesis of the URE2 gene. The two Arg-AGA codons at positions 253 and 254 of the URE2 gene coding sequence were exchanged by CGT codons accordingly.

Differential resistance to proteinase K digestion of the yeast prion-like (Ure2p) protein synthesized in vitro in wheat germ extract and rabbit reticulocyte lysate cell-free translation systems.

The Ure2p yeast prion-like protein was translated in vitro in the presence of labeled [35S]methionine in either rabbit reticulocyte lysate (RRL) or wheat germ extract (WGE) cell-free systems. When subjected to proteinase K digestion, the Ure2p protein synthesized in WGE was proteolysed much more slowly compared to that synthesized in RRL; this displays fragments of about 31-34 kDa, persisting over 8 min. Thus, the digestion rate and pattern of the protein synthesized in WGE, unlike that synthesized in RRL, revealed characteristic features of the [URE3] prion-like isoform of the Ure2p protein [Masison, D.

Construction of a yeast strain deleted for the TRP1 promoter and coding region that enhances the efficiency of the polymerase chain reaction-disruption method.

The sequence of the genome of Saccharomyces cerevisiae was recently determined. As well as all the informations concerning the structure of the chromosomes the scientific community had to deal with the discovery of dozens of new open reading frames (ORFs) of unknown function. The study of these ORFs requires the development of simple procedures that can be used on a large scale.

Functional analysis of YCL09C: evidence for a role as the regulatory subunit of acetolactate synthase.

We have analysed the function of the open reading frame (ORF) YCL09C. The deletion of this ORF from chromosome III does not affect the physiology of the corresponding yeast strain enough to give a distinct phenotype. Nevertheless a computational analysis reveals high homology between this ORF and the enterobacterial genes encoding the regulatory subunit of acetolactate synthase.

Functional domains of fused, a serine-threonine kinase required for signaling in Drosophila.

fused (fu) is a segment-polarity gene encoding a putative serine-threonine kinase. In a wild-type context, all fu mutations display the same set of phenotypes. Nevertheless, mutations of the Suppressor of fused [Su(fu)] gene define three classes of alleles (fuO, fuI, fuII).

Molecular organisation and expression pattern of the segment polarity gene fused of Drosophila melanogaster.

The Drosophila segment-polarity gene fused (fu) is required for pattern formation within embryonic segments and imaginal discs. We previously reported that the 5' part of the fused gene is homologous to the catalytic domain of serine/threonine kinases. We present here the sequence of the complete transcription unit, which predicts a 805 amino acid long protein.

Self-fructification associated with genetic instability in Coprinus radiatus.

In the basidiomycete Coprinus radiatus, crosses gave rise with a high frequency to 2 types of self-fructifying progeny. The first type showed monokaryotic fruiting due to the occurrence of a mutation associated with the B1 incompatibility factor. The second type showed fruiting due to the presence of the 2 parental genomes in the original spore.

Properties of genetic instability during the vegetative growth of Coprinus radiatus.

In Coprinus radiatus, a mutation at the Nic-2 locus unstable at meiosis has been previously described. Further studies have now shown that this mutation is also unstable, although with lower frequency, during vegetative growth. This 'vegetative instability', which is thermosensitive, is not a random process but an aggregative process, perhaps depending on the physiological state of the mycelium.