Heme Degrading Protein HemS Is Involved in Oxidative Stress Response of Bartonella henselae

Bartonellae are hemotropic bacteria, agents of emerging zoonoses. These bacteria are heme auxotroph Alphaproteobacteria which must import heme for supporting their growth, as they cannot synthesize it. Therefore, Bartonella genome encodes for a complete heme uptake system allowing the transportation of this compound across the outer membrane, the periplasm and the inner membranes. Heme has been proposed to be used as an iron source for Bartonella since these bacteria do not synthesize a complete system required for iron Fe3+uptake. Similarly to other bacteria which use heme as an iron source, Bartonellae must transport this compound into the cytoplasm and degrade it to allow the release of iron from the tetrapyrrole ring. For Bartonella, the gene cluster devoted to the synthesis of the complete heme uptake system also contains a gene encoding for a polypeptide that shares homologies with heme trafficking or degrading enzymes. Using complementation of an E. coli mutant strain impaired in heme degradation, we demonstrated that HemS from Bartonella henselae expressed in E. coli allows the release of iron from heme. Purified HemS from B. henselae binds heme and can degrade it in the presence of a suitable electron donor, ascorbate or NADPH-cytochrome P450 reductase. Knocking down the expression of HemS in B. henselae reduces its ability to face H2O2 induced oxidative stress

DprA Is Essential for Natural Competence in Riemerella anatipestifer and Has a Conserved Evolutionary Mechanism

Riemerella anatipestifer ATCC11845 (RA ATCC11845) is naturally competent. However, the genes involved in natural transformation in this species remain largely unknown. Bioinformatic analysis predicts that DprA of RA (DprARa) has three domains: a sterile alpha motif (SAM), a Rossmann fold (RF) domain and a Z-DNA-binding domain (Zα). Inactivation of dprA abrogated natural transformation in RA ATCC11845, and this effect was restored by the expression of dprA in trans. The dprA with SAM and RF domains of Streptococcus pneumoniae and the dprA with RF and Zα domains of Helicobacter pylori was able to restore natural transformation in the RA ATCC11845 dprA mutant. An Arg123 mutation in the RF domain of R. anatipestifer was not able to restore natural transformation of the RA ATCC11845 dprA mutant. Furthermore, DprAR123E abolished its ability to bind DNA, suggesting that the RF domain is essential for the function of DprA. Finally, the dprA of Fusobacterium naviforme which has not been reported to be natural competent currently was partially able to restore natural transformation in RA ATCC11845 dprA mutant. These results collectively suggest that DprA has a conserved evolutionary mechanism

Etude de la regulation de l'expression et de l'activite du facteur de terminaison de transcription rho chez Escherichia coli K 12; recherche des proteines CAP, rho et de l'AMP cyclique chez d'autres bacteries

SIGLECNRS T 56786 / INIST-CNRS - Institut de l'Information Scientifique et TechniqueFRFranc

Functional identification of ygiP as a poitive regulator of the ttdA-ttdB-ygjE operon

International audienceFunctional characterization of unknown genes is currently a major task in biology. The search for gene function involves a combination of various in silico, in vitro and in vivo approaches. Available knowledge from the study of more than 21 LysR-type regulators in Escherichia coli has facilitated the classification of new members of the family. From sequence similarities and its location on the E. coli chromosome, it is suggested that ygiP encodes a lysR regulator controlling the expression of a neighbouring operon; this operon encodes the two subunits of tartrate dehydratase (TtdA, TtdB) and YgiE, an integral inner-membrane protein possibly involved in tartrate uptake. Expression of tartrate dehydratase, which converts tartrate to oxaloacetate, is required for anaerobic growth on glycerol as carbon source in the presence of tartrate. Here, it has been demonstrated that disruption of ygiP, ttdA or ygjE abolishes tartrate-dependent anaerobic growth on glycerol. It has also been shown that tartrate-dependent induction of the ttdA-ttdB-ygjE operon requires a functional YgiP

Mécanisme moléculaire de l'induction de l'expression de l'opéron has de Serratia marcescens

PARIS7-Bibliothèque centrale (751132105) / SudocSudocFranceF

Rôle des protéines paralogues TonB et HasB dans le transport de l'hème par le récepteur de membrane externe HemR de Serratia marcescens

PARIS7-Bibliothèque centrale (751132105) / SudocSudocFranceF

Heme and a Five-Amino-Acid Hemophore Region Form the Bipartite Stimulus Triggering the has Signaling Cascade

Bacterial cells sense the extracellular environment and adapt to that environment by activating gene regulation circuits, often by means of signaling molecules. The Serratia marcescens hemophore is a signaling molecule that acts as an extracellular heme-scavenging protein. The heme-loaded hemophore interacts with its cognate receptor (HasR), triggering transmembrane signaling and turning on transcription of hemophore-dependent heme uptake genes. We investigated the features of the holo-hemophore, the only HasR ligand known to act as an inducer. We used a hemophore mutant that does not deliver its heme and a HasR mutant that does not bind heme, and we showed that heme transfer from the hemophore to the receptor is necessary for induction. Using a hemophore mutant that does not bind heme and that blocks heme transport, we demonstrated that two molecules that do not interact (heme and the mutant hemophore) may nonetheless induce this system. These findings suggest that hemophore-mediated induction and heme transport involve different mechanisms. The hemophore region important for induction was precisely localized to amino acids 50 to 55, which lie in one of the two HasR-binding hemophore regions. This bipartite stimulus probably corresponds to a physiological process because heme is transferred to the receptor before apo-hemophore release. This bipartite regulation mechanism may allow the bacterium to adjust its heme transport mechanism to the perceived environmental heme concentration

HasB, the Serratia marcescens TonB Paralog, Is Specific to HasR▿

Serratia marcescens possesses two functional TonB paralogs, TonBSm and HasB, for energizing TonB-dependent transport receptors (TBDT). Previous work had shown that HasB is specific to heme uptake in the natural host and in Escherichia coli expressing the S. marcescens TBDT receptor HasR, whereas the S. marcescens TonB and E. coli TonB proteins function equally well with various TBDT receptors for heme and siderophores. This has raised the question of the target of this specificity. HasB could be specific either to heme TBDT receptors or only to HasR. To resolve this question, we have cloned in E. coli another S. marcescens heme receptor, HemR, and we show here that this receptor is TonB dependent and does not work with HasB. This demonstrates that HasB is not dedicated to heme TBDT receptors but rather forms a specific pair with HasR. This is the first reported case of a specific TonB protein working with only one TBDT receptor in one given species. We discuss the occurrence, possible molecular mechanisms, and selective advantages of such dedicated TonB paralogs

The Bartonella henselae SitABCD transporter is required for confronting oxidative stress during cell and flea invasion

International audienceBartonella henselae is a zoonotic pathogen that possesses a flea-cat-flea transmission cycle and causes cat scratch disease in humans via cat scratches and bites. In order to establish infection, B. henselae must overcome oxidative stress damage produced by the mammalian host and arthropod vector. B. henselae encodes for putative Fe2+ and Mn2+ transporter SitABCD. In B. henselae, Sitknockdown increases sensitivity to hydrogen peroxide. We consistently show that Sitknockdown decreases the ability of B. henselae to survive in both human endothelial cells and cat fleas, thus demonstrating that the SitABCD transporter plays an important role during the B. henselae infection cycle