"Mais qui es-tu, au juste?" : exploration de l'album Fourchon : entre sentiment d'appartenance et quête d'identité

La séquence didactique proposée en ces pages est à réaliser à partir d’une lecture à voix haute de l’album jeunesse Fourchon de Kyo Maclear, traduit en français par Fanny Britt et illustré par Isabelle Arsenault. Cet album qui aborde de façon originale les différences physiques, psychologiques et culturelles a remporté, en 2012, le Prix jeunesse des libraires du Québec. La séquence qui suit s’adresse particulièrement aux élèves des 2e et 3e cycles du primaire, mais pourrait aussi être exploitée avec des écoliers du 1er cycle, voire avec des élèves du début du secondaire, en adaptant quelque peu certaines activités

From the selfish gene to selfish metabolism: revisiting the central dogma

The standard representation of the Central Dogma (CD) of Molecular Biology conspicuously ignores metabolism. However, both the metabolites and the biochemical fluxes behind any biological phenomenon are encrypted in the DNA sequence. Metabolism constrains and even changes the information flow when the DNA-encoded instructions conflict with the homeostasis of the biochemical network. Inspection of adaptive virulence programs and emergence of xenobiotic-biodegradation pathways in environmental bacteria suggest that their main evolutionary drive is the expansion of their metabolic networks towards new chemical landscapes rather than perpetuation and spreading of their DNA sequences. Faulty enzymatic reactions on suboptimal substrates produce reactive oxygen species (ROS), which fosters DNA diversification and eventually couples catabolism of the new chemicals to growth. All this calls for a revision of the CD in which metabolism (rather than DNA) has the leading role.The work in Author’s laboratory is supported by generous grants of the Spanish Ministry of Science and Innovation (CONSOLIDER), by contracts of the Framework Program of the EU (MICROME, ST-FLOW), the European Research Council (ARISYS) and by Funds of the Autonomous Community of Madrid (PROMT Program).Peer reviewe

The degree of microbiome complexity influences the epithelial response to infection

<p>Abstract</p> <p>Background</p> <p>The human microflora is known to be extremely complex, yet most pathogenesis research is conducted in mono-species models of infection. Consequently, it remains unclear whether the level of complexity of a host's indigenous flora can affect the virulence potential of pathogenic species. Furthermore, it remains unclear whether the colonization by commensal species affects a host cell's response to pathogenic species beyond the direct physical saturation of surface receptors, the sequestration of nutrients, the modulation of the physico-chemical environment in the oral cavity, or the production of bacteriocins. Using oral epithelial cells as a model, we hypothesized that the virulence of pathogenic species may vary depending on the complexity of the flora that interacts with host cells.</p> <p>Results</p> <p>This is the first report that determines the global epithelial transcriptional response to co-culture with defined complex microbiota. In our model, human immortalized gingival keratinocytes (HIGK) were infected with mono- and mixed cultures of commensal and pathogenic species. The global transcriptional response of infected cells was validated and confirmed phenotypically. In our model, commensal species were able to modulate the expression of host genes with a broad diversity of physiological functions and antagonize the effect of pathogenic species at the cellular level. Unexpectedly, the inhibitory effect of commensal species was <it>not </it>correlated with its ability to inhibit adhesion or invasion by pathogenic species.</p> <p>Conclusion</p> <p>Studying the global transcriptome of epithelial cells to single and complex microbial challenges offers clues towards a better understanding of how bacteria-bacteria interactions and bacteria-host interactions impact the overall host response. This work provides evidence that the degree of complexity of a mixed microbiota <it>does </it>influence the transcriptional response to infection of host epithelial cells, and challenges the current dogma regarding the <it>potential </it>versus the <it>actual </it>pathogenicity of bacterial species. These findings support the concept that members of the commensal oral flora have evolved cellular mechanisms that directly modulate the host cell's response to pathogenic species and dampen their relative pathogenicity.</p

Subgingival bacterial colonization profiles correlate with gingival tissue gene expression

Periodontitis is a chronic inflammatory disease caused by the microbiota of the periodontal pocket. We investigated the association between subgingival bacterial profiles and gene expression patterns in gingival tissues of patients with periodontitis. A total of 120 patients undergoing periodontal surgery contributed with a minimum of two interproximal gingival papillae (range 2-4) from a maxillary posterior region. Prior to tissue harvesting, subgingival plaque samples were collected from the mesial and distal aspects of each tissue sample. Gingival tissue RNA was extracted, reverse-transcribed, labeled, and hybridized with whole-genome microarrays (310 in total). Plaque samples were analyzed using checkerboard DNA-DNA hybridizations with respect to 11 bacterial species. Random effects linear regression models considered bacterial levels as exposure and expression profiles as outcome variables. Gene Ontology analyses summarized the expression patterns into biologically relevant categories. Wide inter-species variation was noted in the number of differentially expressed gingival tissue genes according to subgingival bacterial levels: Using a Bonferroni correction (p < 9.15 × 10-7), 9,392 probe sets were differentially associated with levels of Tannerella forsythia, 8,537 with Porphyromonas gingivalis, 6,460 with Aggregatibacter actinomycetemcomitans, 506 with Eikenella corrodens and only 8 with Actinomyces naeslundii. Cluster analysis identified commonalities and differences among tissue gene expression patterns differentially regulated according to bacterial levels. Our findings suggest that the microbial content of the periodontal pocket is a determinant of gene expression in the gingival tissues and provide new insights into the differential ability of periodontal species to elicit a local host response

Production of human entorhinal stellate cell-like cells by forward programming shows an important role of Foxp1 in reprogramming

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Identification of Leishmania Proteins Preferentially Released in Infected Cells Using Change Mediated Antigen Technology (CMAT)

Although Leishmania parasites have been shown to modulate their host cell's responses to multiple stimuli, there is limited evidence that parasite molecules are released into infected cells. In this study, we present an implementation of the change mediated antigen technology (CMAT) to identify parasite molecules that are preferentially expressed in infected cells. Sera from mice immunized with cell lysates prepared from L. donovani or L. pifanoi-infected macrophages were adsorbed with lysates of axenically grown amastigotes of L. donovani or L. pifanoi, respectively, as well as uninfected macrophages. The sera were then used to screen inducible parasite expression libraries constructed with genomic DNA. Eleven clones from the L. pifanoi and the L. donovani screen were selected to evaluate the characteristics of the molecules identified by this approach. The CMAT screen identified genes whose homologs encode molecules with unknown function as well as genes that had previously been shown to be preferentially expressed in the amastigote form of the parasite. In addition a variant of Tryparedoxin peroxidase that is preferentially expressed within infected cells was identified. Antisera that were then raised to recombinant products of the clones were used to validate that the endogenous molecules are preferentially expressed in infected cells. Evaluation of the distribution of the endogenous molecules in infected cells showed that some of these molecules are secreted into parasitophorous vacuoles (PVs) and that they then traffic out of PVs in vesicles with distinct morphologies. This study is a proof of concept study that the CMAT approach can be applied to identify putative Leishmania parasite effectors molecules that are preferentially expressed in infected cells. In addition we provide evidence that Leishmania molecules traffic out of the PV into the host cell cytosol and nucleus

Application of In Vivo Induced Antigen Technology (IVIAT) to Bacillus anthracis

In vivo induced antigen technology (IVIAT) is an immuno-screening technique that identifies bacterial antigens expressed during infection and not during standard in vitro culturing conditions. We applied IVIAT to Bacillus anthracis and identified PagA, seven members of a N-acetylmuramoyl-L-alanine amidase autolysin family, three P60 family lipoproteins, two transporters, spore cortex lytic protein SleB, a penicillin binding protein, a putative prophage holin, respiratory nitrate reductase NarG, and three proteins of unknown function. Using quantitative real-time PCR comparing RNA isolated from in vitro cultured B. anthracis to RNA isolated from BALB/c mice infected with virulent Ames strain B. anthracis, we confirmed induced expression in vivo for a subset of B. anthracis genes identified by IVIAT, including L-alanine amidases BA3767, BA4073, and amiA (pXO2-42); the bacteriophage holin gene BA4074; and pagA (pXO1-110). The exogenous addition of two purified putative autolysins identified by IVIAT, N-acetylmuramoyl-L-alanine amidases BA0485 and BA2446, to vegetative B. anthracis cell suspensions induced a species-specific change in bacterial morphology and reduction in viable bacterial cells. Many of the proteins identified in our screen are predicted to affect peptidoglycan re-modeling, and our results support significant cell wall structural remodeling activity during B. anthracis infection. Identification of L-alanine amidases with B. anthracis specificity may suggest new potential therapeutic targets

Single-cell Atlas of common variable immunodeficiency shows germinal center-associated epigenetic dysregulation in B-cell responses.

Common variable immunodeficiency (CVID), the most prevalent symptomatic primary immunodeficiency, displays impaired terminal B-cell differentiation and defective antibody responses. Incomplete genetic penetrance and ample phenotypic expressivity in CVID suggest the participation of additional pathogenic mechanisms. Monozygotic (MZ) twins discordant for CVID are uniquely valuable for studying the contribution of epigenetics to the disease. Here, we generate a single-cell epigenomics and transcriptomics census of naïve-to-memory B cell differentiation in a CVID-discordant MZ twin pair. Our analysis identifies DNA methylation, chromatin accessibility and transcriptional defects in memory B-cells mirroring defective cell-cell communication upon activation. These findings are validated in a cohort of CVID patients and healthy donors. Our findings provide a comprehensive multi-omics map of alterations in naïve-to-memory B-cell transition in CVID and indicate links between the epigenome and immune cell cross-talk. Our resource, publicly available at the Human Cell Atlas, gives insight into future diagnosis and treatments of CVID patients.We thank the CERCA Program/Generalitat de Catalunya and the Josep Carreras Foundation for institutional support. This publication is part of the Human Cell Atlas; www.humancellatlas.org/publications. This study was funded by: Spanish Ministry of Science and Innovation (grant number PID2020-117212RB-I00/AEI/10.13038/501100011033) (E.B.), Instituto de Salud Carlos III (ISCIII), Ref. AC18/00057, associated with i-PAD project (ERARE European Union program) (E.B.), the Jeffrey Modell Foundation (E.B.), Wellcome Sanger core funding (grant no. WT206194) (R.V.-T.), the Chan Zuckerberg Initiative (grant 2020-216799) (R.V.-T. and E.B.), an EMBO short-term fellowship (J.R.U.), Fondo de Investigación Sanitaria Instituto de Salud Carlos III (FIS PI16/01605) (L.P.-M.), the Spanish Ministry of Science, Innovation and Universities (SAF2017-89109-P; AEI/FEDER, UE) (H.H.), Instituto de Salud Carlos III, Ministry of Health (PI16/00759) and European Regional Development Fund-European Social Fund—FEDER-FSE) (C.R-G.), Grupo DISA (OA18/017) (C.R.-G.), the UK Biotechnology and Biological Sciences Research Council (BBS/E/B/000C0426) (G.K.) and Medical Research Council (MR/S000437/1) (G.K.). We are indebted to the donors for participating in this research. We thank Antonio Garcia-Gomez for graphical design support, Sarah Teichmann for her useful feedback, Hamish King for helping with single-cell germinal center dataset availability, Xi Chen for performing scATAC-seq analysis, Kirsty Ambridge and Elena Prigmore for their involvement in single-cell RNA library generation, Martin Prete for creating online visualizations for our cell atlas and Esther Castaño and Beatriz Barroso from CCiTUB Cytometry Unit for their support with single-cell sorting and Dr. Carla Gianelli and Dr. Rebeca Rodríguez Pena for the patient follow-up in the CVID cohort

Alanine Racemase Mutants of Burkholderia pseudomallei and Burkholderia mallei and Use of Alanine Racemase as a Non-Antibiotic-Based Selectable Marker

Burkholderia pseudomallei and Burkholderia mallei are category B select agents and must be studied under BSL3 containment in the United States. They are typically resistant to multiple antibiotics, and the antibiotics used to treat B. pseudomallei or B. mallei infections may not be used as selective agents with the corresponding Burkholderia species. Here, we investigated alanine racemase deficient mutants of B. pseudomallei and B. mallei for development of non-antibiotic-based genetic selection methods and for attenuation of virulence. The genome of B. pseudomallei K96243 has two annotated alanine racemase genes (bpsl2179 and bpss0711), and B. mallei ATCC 23344 has one (bma1575). Each of these genes encodes a functional enzyme that can complement the alanine racemase deficiency of Escherichia coli strain ALA1. Herein, we show that B. pseudomallei with in-frame deletions in both bpsl2179 and bpss0711, or B. mallei with an in-frame deletion in bma1575, requires exogenous d-alanine for growth. Introduction of bpsl2179 on a multicopy plasmid into alanine racemase deficient variants of either Burkholderia species eliminated the requirement for d-alanine. During log phase growth without d-alanine, the viable counts of alanine racemase deficient mutants of B. pseudomallei and B. mallei decreased within 2 hours by about 1000-fold and 10-fold, respectively, and no viable bacteria were present at 24 hours. We constructed several genetic tools with bpsl2179 as a selectable genetic marker, and we used them without any antibiotic selection to construct an in-frame ΔflgK mutant in the alanine racemase deficient variant of B. pseudomallei K96243. In murine peritoneal macrophages, wild type B. mallei ATCC 23344 was killed much more rapidly than wild type B. pseudomallei K96243. In addition, the alanine racemase deficient mutant of B. pseudomallei K96243 exhibited attenuation versus its isogenic parental strain with respect to growth and survival in murine peritoneal macrophages