Biblioteca Digital

Implantation of the fertilized egg into the maternal uterus depends on the fine balance between inflammatory and anti-inflammatory processes. Whilst regulatory T cells (Tregs) are reportedly involved in protection of allogeneic fetuses against rejection by the maternal immune system, their role for pregnancy to establish, e.g., blastocyst implantation, is not clear. By using 2-photon imaging we show that Foxp3(+) cells accumulated in the mouse uterus during the receptive phase of the estrus cycle. Seminal fluid further fostered Treg expansion. Depletion of Tregs in two Foxp3.DTR-based models prior to pairing drastically impaired implantation and resulted in infiltration of activated T effector cells as well as in uterine inflammation and fibrosis in both allogeneic and syngeneic mating combinations. Genetic deletion of the homing receptor CCR7 interfered with accumulation of Tregs in the uterus and implantation indicating that homing of Tregs to the uterus was mediated by CCR7. Our results demonstrate that Tregs play a critical role in embryo implantation by preventing the development of a hostile uterine microenvironment.; DFG grants: (ZE526/4-2, SFB854TP7), Wilhelm Sander Stiftung Germany grant: (2009.022.1), Helmholtz Alliance for Immunotherapy...

Hundreds of genetic variants have been associated to common diseases through genome-wide association studies (GWAS), yet there are limits to current approaches in detecting true small effect risk variants against a background of false positive findings. Here we addressed the missing heritability problem, aiming to test whether there are indeed risk variants within GWAS statistical noise and to develop a systematic strategy to retrieve these hidden variants. Employing an integrative approach, which combines protein-protein interactions with association data from GWAS for 6 common diseases, we found that associated-genes at less stringent significance levels (p < 0.1) with any of these diseases are functionally connected beyond noise expectation. This functional coherence was used to identify disease-relevant subnetworks, which were shown to be enriched in known genes, outperforming the selection of top GWAS genes. As a proof of principle, we applied this approach to breast cancer, supporting well-known breast cancer genes, while pinpointing novel susceptibility genes for experimental validation. This study reinforces the idea that GWAS are under-analyzed and that missing heritability is rather hidden. It extends the use of protein networks to reveal this missing heritability...

Plasmodium falciparum malaria in India is characterized by high rates of severe disease, with multiple organ dysfunction (MOD)-mainly associated with acute renal failure (ARF)-and increased mortality. The objective of this study is to identify cytokine signatures differentiating severe malaria patients with MOD, cerebral malaria (CM), and cerebral malaria with MOD (CM-MOD) in India. We have previously shown that two cytokines clusters differentiated CM from mild malaria in Maharashtra. Hence, we also aimed to determine if these cytokines could discriminate malaria subphenotypes in Odisha.; Indo-French Centre for the Promotion of Advanced Research (IFCPAR project No 3703), International Associated Laboratory Systems Immunology and Genetics of Infectious Diseases (LIA SIGID, CNRS, Lille University and the Department of Biotechnology (DBT), Ministry of Science and Technology of India and the ANR LAbEx Parafrap).

Co-evolution between the mammalian immune system and the gut microbiota is believed to have shaped the microbiota's astonishing diversity. Here we test the corollary hypothesis that the adaptive immune system, directly or indirectly, influences the evolution of commensal species. We compare the evolution of Escherichia coli upon colonization of the gut of wild-type and Rag2(-/-) mice, which lack lymphocytes. We show that bacterial adaptation is slower in immune-compromised animals, a phenomenon explained by differences in the action of natural selection within each host. Emerging mutations exhibit strong beneficial effects in healthy hosts but substantial antagonistic pleiotropy in immune-deficient mice. This feature is due to changes in the composition of the gut microbiota, which differs according to the immune status of the host. Our results indicate that the adaptive immune system influences the tempo and predictability of E. coli adaptation to the mouse gut.; EU-FP7 infrastructure grants: (EMMA and InfrafrontierI3), LAO/ITQB, FCT grant: (SFRH/BD/80257/2011).

Muller's ratchet is an evolutionary process that has been implicated in the extinction of asexual species, the evolution of non-recombining genomes, such as the mitochondria, the degeneration of the Y chromosome, and the evolution of sex and recombination. Here we study the speed of Muller's ratchet in a spatially structured population which is subdivided into many small populations (demes) connected by migration, and distributed on a graph. We studied different types of networks: regular networks (similar to the stepping-stone model), small-world networks and completely random graphs. We show that at the onset of the small-world network - which is characterized by high local connectivity among the demes but low average path length - the speed of the ratchet starts to decrease dramatically. This result is independent of the number of demes considered, but is more pronounced the larger the network and the stronger the deleterious effect of mutations. Furthermore, although the ratchet slows down with increasing migration between demes, the observed decrease in speed is smaller in the stepping-stone model than in small-world networks. As migration rate increases, the structured populations approach, but never reach, the result in the corresponding panmictic population with the same number of individuals. Since small-world networks have been shown to describe well the real contact networks among people...

Muller's ratchet is an evolutionary process that has been implicated in the extinction of asexual species, the evolution of mitochondria, the degeneration of the Y chromosome, the evolution of sex and recombination and the evolution of microbes. Here we study the speed of Muller's ratchet in a population subdivided into many small subpopulations connected by migration, and distributed on a network. We compare the speed of the ratchet in two distinct types of topologies: scale free networks and random graphs. The difference between the topologies is noticeable when the average connectivity of the network and the migration rate is large. In this situation we observe that the ratchet clicks faster in scale free networks than in random graphs. So contrary to intuition, scale free networks are more prone to loss of genetic information than random graphs. On the other hand, we show that scale free networks are more robust to the random extinction than random graphs. Since these complex networks have been shown to describe well real-life systems, our results open a framework for studying the evolution of microbes and disease epidemics.

The levels and patterns of variation at a neutral locus are analyzed in a haploid asexual population Undergoing accumulation of deleterious mutations due to Muller's ratchet. We find that the movement Of Muller's ratchet can be associated with a considerable reduction in genetic diversity below classical neutral expectation. The extent to which variability is reduced is a function Of the deleterious initiation rate, the fitness effects of the Imitations, and the population size. Approximate analytical expressions for the expected genetic diversity are compared with simulation results under two different models of deleterious imitations: a model where all deleterious imitations have equal effects and a model where there are two classes of deleterious imitations. We also find that Muller's ratchet can produce a considerable distortion in the neutral frequency spectrum toward an excess of rare variants.

Deriving useful microsatellite markers in lepidopterans has been challenging when relying on scans of genomic DNA libraries, presumably due to repetitiveness in their genomes. We assayed 96 of 320 microsatellites identified in silico from a collection of Bicyclus anynana ESTs, in 11 independent individuals from a laboratory population.From the 68 successful assays, we identified 40 polymorphic markers including 22 with BLAST-based annotation. Nine of 12 selected polymorphic markers tested in a panel of 24 wild-caught individuals converted to successful assays and were all polymorphic. We discuss how microsatellite discovery in ESTs is an efficient strategy with important attendant advantages.

Recent public health threats have propelled major innovations on infectious disease monitoring, culminating in the development of innovative syndromic surveillance methods. Influenzanet is an internet-based system that monitors influenza-like illness (ILI) in cohorts of self-reporting volunteers in European countries since 2003. We investigate and confirm coherence through the first ten years in comparison with ILI data from the European Influenza Surveillance Network and demonstrate country-specific behaviour of participants with ILI regarding medical care seeking. Using regression analysis, we determine that chronic diseases, being a child, living with children, being female, smoking and pets at home, are all independent predictors of ILI risk, whereas practicing sports and walking or bicycling for locomotion are associated with a small risk reduction. No effect for using public transportation or living alone was found. Furthermore, we determine the vaccine effectiveness for ILI for each season.; Institute for the Promotion of Innovation by Science and Technology (IWT - strategic basic research project SIMID), ICyT projects: (60/2010, 343/2010), Brazilian Research Council (CNPq), CAPES.

The hypoxia-inducible factor (HIF) is a key regulator of the cellular response to hypoxia which promotes oxygen delivery and metabolic adaptation to oxygen deprivation. However, the degree and duration of HIF-1α expression in hypoxia must be carefully balanced within cells in order to avoid unwanted side effects associated with excessive activity. The expression of HIF-1α mRNA is suppressed in prolonged hypoxia, suggesting that the control of HIF1A gene transcription is tightly regulated by negative feedback mechanisms. Little is known about the resolution of the HIF-1α protein response and the suppression of HIF-1α mRNA in prolonged hypoxia. Here, we demonstrate that the Repressor Element 1-Silencing Transcription factor (REST) binds to the HIF-1α promoter in a hypoxia-dependent manner. Knockdown of REST using RNAi increases the expression of HIF-1α mRNA, protein and transcriptional activity. Furthermore REST knockdown increases glucose consumption and lactate production in a HIF-1α- (but not HIF-2α-) dependent manner. Finally, REST promotes the resolution of HIF-1α protein expression in prolonged hypoxia. In conclusion, we hypothesize that REST represses transcription of HIF-1α in prolonged hypoxia, thus contributing to the resolution of the HIF-1α response.; Science Foundation Ireland grant numnber: (06/CE/B1129).

Activation of NF-κB-dependent transcription represents an important hallmark of inflammation. While the acute inflammatory response is per se beneficial, it can become deleterious if its spatial and temporal profile is not tightly controlled. Classically, NF-κB activity is limited by cytoplasmic retention of the NF-κB dimer through binding to inhibitory IκB proteins. However, increasing evidence suggests that NF-κB activity can also be efficiently contained by direct ubiquitination of NF-κB subunits. Here, we identify the HECT-domain ubiquitin ligase HERC3 as novel negative regulator of NF-κB activity. We find that HERC3 restricts NF-κB nuclear import and DNA binding without affecting IκBα degradation. Instead HERC3 indirectly binds to the NF-κB RelA subunit after liberation from IκBα inhibitor leading to its ubiquitination and protein destabilization. Remarkably, the regulation of RelA activity by HERC3 is independent of its inherent ubiquitin ligase activity. Rather, we show that HERC3 and RelA are part of a multi-protein complex containing the proteasome as well as the ubiquitin-like protein ubiquilin-1 (UBQLN1). We present evidence that HERC3 and UBQLN1 provide a link between NF-κB RelA and the 26S proteasome, thereby facilitating RelA protein degradation. Our findings establish HERC3 as novel candidate regulating the inflammatory response initiated by NF-κB.; American Heart Association Scientist Development: (SDG102600298)...

Symbiotic interactions between microbes and their multicellular hosts have manifold impacts on molecular, cellular and organismal biology. To identify candidate bacterial genes involved in maintaining endosymbiotic associations with insect hosts, we analyzed genome-wide patterns of gene expression in the alpha-proteobacteria Wolbachia pipientis across the life cycle of Drosophila melanogaster using public data from the modENCODE project that was generated in a Wolbachia-infected version of the ISO1 reference strain. We find that the majority of Wolbachia genes are expressed at detectable levels in D. melanogaster across the entire life cycle, but that only 7.8% of 1195 Wolbachia genes exhibit robust stage- or sex-specific expression differences when studied in the "holo-organism" context. Wolbachia genes that are differentially expressed during development are typically up-regulated after D. melanogaster embryogenesis, and include many bacterial membrane, secretion system and ankyrin-repeat containing proteins. Sex-biased genes are often organised as small operons of uncharacterised genes and are mainly up-regulated in adult males D. melanogaster in an age-dependent manner suggesting a potential role in cytoplasmic incompatibility. Our results indicate that large changes in Wolbachia gene expression across the Drosophila life-cycle are relatively rare when assayed across all host tissues...

Rheumatoid arthritis (RA) is a chronic immune-mediated inflammatory disease characterized by cellular infiltration into the joints, hyperproliferation of synovial cells and bone damage. Available treatments for RA only induce remission in around 30% of the patients, have important adverse effects and its use is limited by their high cost. Therefore, compounds that can control arthritis, with an acceptable safety profile and low production costs are still an unmet need. We have shown, in vitro, that celastrol inhibits both IL-1β and TNF, which play an important role in RA, and, in vivo, that celastrol has significant anti-inflammatory properties. Our main goal in this work was to test the effect of celastrol in the number of sublining CD68 macrophages (a biomarker of therapeutic response for novel RA treatments) and on the overall synovial tissue cellularity and joint structure in the adjuvant-induced rat model of arthritis (AIA).; FCT fellowship: (SFRH/BPD/92860/2013).

Visceral leishmaniasis is a severe and potentially fatal disease caused by protozoa of the genus Leishmania, transmitted by phlebotomine sandflies. In Europe and the Mediterranean region, L. infantum is the commonest agent of visceral leishmaniasis, causing a wide spectrum of clinical manifestations, including asymptomatic carriage, cutaneous lesions and severe visceral disease. Visceral leishmaniasis is more frequent in immunocompromised individuals and data obtained in experimental models of infection have highlighted the importance of the host immune response, namely the efficient activation of host's macrophages, in determining infection outcome. Conversely, few studies have addressed a possible contribution of parasite variability to this outcome.; No funders or funding refered in the paper.

LAO/ITQB, FCT.

Populations facing novel environments are expected to evolve through the accumulation of adaptive substitutions. The dynamics of adaptation depend on the fitness landscape and possibly on the genetic background on which new mutations arise. Here, we model the dynamics of adaptive evolution at the phenotypic and genotypic levels, focusing on a Fisherian landscape characterized by a single peak. We find that Fisher's geometrical model of adaptation, extended to allow for small random environmental variations, is able to explain several features made recently in experimentally evolved populations. Consistent with data on populations evolving under controlled conditions, the model predicts that mean population fitness increases rapidly when populations face novel environments and then achieves a dynamic plateau, the rate of molecular evolution is remarkably constant over long periods of evolution, mutators are expected to invade and patterns of epistasis vary along the adaptive walk. Negative epistasis is expected in the initial steps of adaptation but not at later steps, a prediction that remains to be tested. Furthermore, populations are expected to exhibit high levels of phenotypic diversity at all times during their evolution. This implies that populations are possibly able to adapt rapidly to novel abiotic environments.; CAPES-IGC.

The genomes of species of Escherichia coli (E. coli) show an extraordinary amount of diversity, which include commensal strains and strains belonging to different pathovars. Many strains of E. coli, which can cause mild or severe pathologies in humans, have a commensal ancestor. Understanding the evolutionary changes that can lead to a transition from commensal to pathogen is an important task, which requires integration of different methodologies. One method is experimental evolution of bacteria, in controlled environments, that mimic some of the selective pressures, likely to be important during the transition to pathogenesis. The success of such a transition will depend, at least partially, on ability of E. coli to adapt to the presence of cells of the immune system. Here, we describe a protocol for performing experimental evolution of a commensal strain of E. coli, a derivative of the well studied K12, under the constant selective pressure imposed by cells of the innate immune system, specifically RAW 264.7 murine macrophage cell line.; LAO/ITQB, FCT.

Unraveling the factors that determine the rate of adaptation is a major question in evolutionary biology. One key parameter is the effect of a new mutation on fitness, which invariably depends on the environment and genetic background. The fate of a mutation also depends on population size, which determines the amount of drift it will experience. Here, we manipulate both population size and genotype composition and follow adaptation of 23 distinct Escherichia coli genotypes. These have previously accumulated mutations under intense genetic drift and encompass a substantial fitness variation. A simple rule is uncovered: the net fitness change is negatively correlated with the fitness of the genotype in which new mutations appear--a signature of epistasis. We find that Fisher's geometrical model can account for the observed patterns of fitness change and infer the parameters of this model that best fit the data, using Approximate Bayesian Computation. We estimate a genomic mutation rate of 0.01 per generation for fitness altering mutations, albeit with a large confidence interval, a mean fitness effect of mutations of -0.01, and an effective number of traits nine in mutS(-) E. coli. This framework can be extended to confront a broader range of models with data and test different classes of fitness landscape models.; LAO/ITQB...

Ohno's hypothesis states that dosage compensation in mammals evolved in two steps: a twofold hyperactivation of the X chromosome in both sexes to compensate for gene losses on the Y chromosome, and silencing of one X (X-chromosome inactivation, XCI) in females to restore optimal dosage. Recent tests of this hypothesis have returned contradictory results. In this review, we explain this ongoing controversy and argue that a novel view on dosage compensation evolution in mammals is starting to emerge. Ohno's hypothesis may be true for a few, dosage-sensitive genes only. If so few genes are compensated, then why has XCI evolved as a chromosome-wide mechanism? This and several other questions raised by the new data in mammals are discussed, and future research directions are proposed.; Agence Nationale de la Recherche grant number: (ANR-12-BSV7-0002), Instituto Gulbenkian de Ciência.

The evolutionary role of transposable elements (TEs) is still highly controversial. Two key parameters, the transposition rate (u and w, for replicative and non-replicative transposition) and the excision rate (e) are fundamental to understanding their evolution and maintenance in populations. We have estimated u, w and e for six families of TEs (including eight members: IS1, IS2, IS3, IS4, IS5, IS30, IS150 and IS186) in Escherichia coli, using a mutation accumulation (MA) experiment. In this experiment, mutations accumulate essentially at the rate at which they appear, during a period of 80 500 (1610 generations × 50 lines) generations, and spontaneous transposition events can be detected. This differs from other experiments in which insertions accumulated under strong selective pressure or over a limited genomic target. We therefore provide new estimates for the spontaneous rates of transposition and excision in E. coli. We observed 25 transposition and three excision events in 50 MA lines, leading to overall rate estimates of u ∼ 1.15 × 10(-5), w ∼ 4 × 10(-8) and e ∼ 1.08 × 10(-6) (per element, per generation). Furthermore, extensive variation between elements was found, consistent with previous knowledge of the mechanisms and regulation of transposition for the different elements.; Natural Sciences and Engineering Research Council of Canada...