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- Microbial Diversity and Toxin Risk in Tropical Freshwater Reservoirs of Cape VerdePublication . Semedo-Aguiar, Ana; Pereira-Leal, Jose; Leite, RicardoThe Cape Verde islands are part of the African Sahelian arid belt that possesses an erratic rain pattern prompting the need for water reservoirs, which are now critical for the country’s sustainability. Worldwide, freshwater cyanobacterial blooms are increasing in frequency due to global climate change and the eutrophication of water bodies, particularly in reservoirs. To date, there have been no risk assessments of cyanobacterial toxin production in these man-made structures. We evaluated this potential risk using 16S rRNA gene amplicon sequencing and full metagenome sequencing in freshwater reservoirs of Cape Verde. Our analysis revealed the presence of several potentially toxic cyanobacterial genera in all sampled reservoirs. Faveta potentially toxic and bloom-forming Microcystis sp., dominated our samples, while a Cryptomonas green algae and Gammaproteobacteria dominated Saquinho and Poilão reservoirs. We reconstructed and assembled the Microcystis genome, extracted from the metagenome of bulk DNA from Faveta water. Phylogenetic analysis of Microcystis cf. aeruginosa CV01’s genome revealed its close relationship with other Microcystis genomes, as well as clustering with other continental African strains, suggesting geographical coherency. In addition, it revealed several clusters of known toxin-producing genes. This survey reinforces the need to better understand the country’s microbial ecology as a whole of water reservoirs on the rise.
- Collective electrical oscillations of a diatom population induced by dark stressPublication . Rocha, Paulo R. F.; Silva, Alexandra D.; Godinho, Lia; Dane, Willem; Estrela, Pedro; Vandamme, Lode K. J.; Pereira-Leal, Jose B.; de Leeuw, Dago M.; Leite, Ricardo B.Diatoms are photosynthetic microalgae, a group with a major environmental role on the planet due to the biogeochemical cycling of silica and global fixation of carbon. However, they can evolve into harmful blooms through a resourceful communication mechanism, not yet fully understood. Here, we demonstrate that a population of diatoms under darkness show quasi-periodic electrical oscillations, or intercellular waves. The origin is paracrine signaling, which is a feedback, or survival, mechanism that counteracts changes in the physicochemical environment. The intracellular messenger is related to Ca2+ions since spatiotemporal changes in their concentration match the characteristics of the intercellular waves. Our conclusion is supported by using a Ca2+channel inhibitor. The transport of Ca2+ions through the membrane to the extracellular medium is blocked and the intercellular waves disappear. The translation of microalgae cooperative signaling paves the way for early detection and prevention of harmful blooms and an extensive range of stress-induced alterations in the aquatic ecosystem.
- Genetic Competence Drives Genome Diversity in Bacillus subtilisPublication . Brito, Patrícia H; Chevreux, Bastien; Serra, Cláudia R; Schyns, Ghislain; Henriques, Adriano O; Pereira-Leal, José BProkaryote genomes are the result of a dynamic flux of genes, with increases achieved via horizontal gene transfer and reductions occurring through gene loss. The ecological and selective forces that drive this genomic flexibility vary across species. Bacillus subtilis is a naturally competent bacterium that occupies various environments, including plant-associated, soil, and marine niches, and the gut of both invertebrates and vertebrates. Here, we quantify the genomic diversity of B. subtilis and infer the genome dynamics that explain the high genetic and phenotypic diversity observed. Phylogenomic and comparative genomic analyses of 42 B. subtilis genomes uncover a remarkable genome diversity that translates into a core genome of 1,659 genes and an asymptotic pangenome growth rate of 57 new genes per new genome added. This diversity is due to a large proportion of low-frequency genes that are acquired from closely related species. We find no gene-loss bias among wild isolates, which explains why the cloud genome, 43% of the species pangenome, represents only a small proportion of each genome. We show that B. subtilis can acquire xenologous copies of core genes that propagate laterally among strains within a niche. While not excluding the contributions of other mechanisms, our results strongly suggest a process of gene acquisition that is largely driven by competence, where the long-term maintenance of acquired genes depends on local and global fitness effects. This competence-driven genomic diversity provides B. subtilis with its generalist character, enabling it to occupy a wide range of ecological niches and cycle through them.
- Actin stress fiber organization promotes cell stiffening and proliferation of pre-invasive breast cancer cellsPublication . Tavares, Sandra; Vieira, André Filipe; Taubenberger, Anna Verena; Araújo, Margarida; Martins, Nuno Pimpao; Brás-Pereira, Catarina; Polónia, António; Herbig, Maik; Barreto, Clara; Otto, Oliver; Cardoso, Joana; Pereira-Leal, José B.; Guck, Jochen; Paredes, Joana; Janody, FlorenceStudies of the role of actin in tumour progression have highlighted its key contribution in cell softening associated with cell invasion. Here, using a human breast cell line with conditional Src induction, we demonstrate that cells undergo a stiffening state prior to acquiring malignant features. This state is characterized by the transient accumulation of stress fibres and upregulation of Ena/VASP-like (EVL). EVL, in turn, organizes stress fibres leading to transient cell stiffening, ERK-dependent cell proliferation, as well as enhancement of Src activation and progression towards a fully transformed state. Accordingly, EVL accumulates predominantly in premalignant breast lesions and is required for Src-induced epithelial overgrowth in Drosophila. While cell softening allows for cancer cell invasion, our work reveals that stress fibre-mediated cell stiffening could drive tumour growth during premalignant stages. A careful consideration of the mechanical properties of tumour cells could therefore offer new avenues of exploration when designing cancer-targeting therapies.
- RAG Recombinase as a Selective Pressure for Genome EvolutionPublication . Passagem-Santos, D.; Bonnet, M.; Sobral, D.; Trancoso, I.; Silva, J.G.; Barreto, V.M.; Athanasiadis, A.; Demengeot, J.; Pereira-Leal, J.B.The RAG recombinase is a domesticated transposable element co-opted in jawed vertebrates to drive the process of the so-called V(D)J recombination, which is the hallmark of the adaptive immune system to produce antigen receptors. RAG targets, namely, the Recombination Signal Sequences (RSS), are rather long and degenerated sequences, which highlights the ability of the recombinase to interact with a wide range of target sequences, including outside of antigen receptor loci. The recognition of such cryptic targets by the recombinase threatens genome integrity by promoting aberrant DNA recombination, as observed in lymphoid malignancies. Genomes evolution resulting from RAG acquisition is an ongoing discussion, in particular regarding the counter-selection of sequences resembling the RSS and the modifications of epigenetic regulation at these potential cryptic sites. Here, we describe a new bioinformatics tool to map potential RAG targets in all jawed vertebrates. We show that our REcombination Classifier (REC) outperforms the currently available tool and is suitable for full genomes scans from species other than human and mouse. Using the REC, we document a reduction in density of potential RAG targets at the transcription start sites of genes co-expressed with the rag genes and marked with high levels of the trimethylation of the lysine 4 of the histone 3 (H3K4me3), which correlates with the retention of functional RAG activity after the horizontal transfer.
- CYR61 and TAZ Upregulation and Focal Epithelial to Mesenchymal Transition May Be Early Predictors of Barrett’s Esophagus Malignant ProgressionPublication . Cardoso, Joana; Mesquita, Marta; Dias Pereira, António; Bettencourt-Dias, Mónica; Chaves, Paula; Pereira-Leal, José B.Barrett's esophagus is the major risk factor for esophageal adenocarcinoma. It has a low but non-neglectable risk, high surveillance costs and no reliable risk stratification markers. We sought to identify early biomarkers, predictive of Barrett's malignant progression, using a meta-analysis approach on gene expression data. This in silico strategy was followed by experimental validation in a cohort of patients with extended follow up from the Instituto Português de Oncologia de Lisboa de Francisco Gentil EPE (Portugal). Bioinformatics and systems biology approaches singled out two candidate predictive markers for Barrett's progression, CYR61 and TAZ. Although previously implicated in other malignancies and in epithelial-to-mesenchymal transition phenotypes, our experimental validation shows for the first time that CYR61 and TAZ have the potential to be predictive biomarkers for cancer progression. Experimental validation by reverse transcriptase quantitative PCR and immunohistochemistry confirmed the up-regulation of both genes in Barrett's samples associated with high-grade dysplasia/adenocarcinoma. In our cohort CYR61 and TAZ up-regulation ranged from one to ten years prior to progression to adenocarcinoma in Barrett's esophagus index samples. Finally, we found that CYR61 and TAZ over-expression is correlated with early focal signs of epithelial to mesenchymal transition. Our results highlight both CYR61 and TAZ genes as potential predictive biomarkers for stratification of the risk for development of adenocarcinoma and suggest a potential mechanistic route for Barrett's esophagus neoplastic progression.
- inTB - a data integration platform for molecular and clinical epidemiological analysis of tuberculosisPublication . Soares, Patrícia; Alves, Renato J; Abecasis, Ana B; Penha-Gonçalves, Carlos; Gomes, M Gabriela M; Pereira-Leal, José BTuberculosis is currently the second highest cause of death from infectious diseases worldwide. The emergence of multi and extensive drug resistance is threatening to make tuberculosis incurable. There is growing evidence that the genetic diversity of Mycobacterium tuberculosis may have important clinical consequences. Therefore, combining genetic, clinical and socio-demographic data is critical to understand the epidemiology of this infectious disease, and how virulence and other phenotypic traits evolve over time. This requires dedicated bioinformatics platforms, capable of integrating and enabling analyses of this heterogeneous data.
- Genome of a Gut Strain of Bacillus subtilisPublication . Schyns, G.; Serra, C. R.; Lapointe, T.; Pereira-Leal, J. B.; Potot, S.; Fickers, P.; Perkins, J. B.; Wyss, M.; Henriques, A. O.Bacillus subtilis is a Gram-positive, rod-shaped, spore-forming bacterium. We present the genome sequence of an undomesticated strain, BSP1, isolated from poultry. The sequence of the BSP1 genome supports the view that B. subtilis has a biphasic lifestyle, cycling between the soil and the animal gastrointestinal tract, and it provides molecular-level insight into the adaptation of B. subtilis to life under laboratory conditions.
- Aurora at the pole and equator: overlapping functions of Aurora kinases in the mitotic spindlePublication . Hochegger, H.; Hegarat, N.; Pereira-Leal, J. B.The correct assembly and timely disassembly of the mitotic spindle is crucial for the propagation of the genome during cell division. Aurora kinases play a central role in orchestrating bipolar spindle establishment, chromosome alignment and segregation. In most eukaryotes, ranging from amoebas to humans, Aurora activity appears to be required both at the spindle pole and the kinetochore, and these activities are often split between two different Aurora paralogues, termed Aurora A and B. Polar and equatorial functions of Aurora kinases have generally been considered separately, with Aurora A being mostly involved in centrosome dynamics, whereas Aurora B coordinates kinetochore attachment and cytokinesis. However, double inactivation of both Aurora A and B results in a dramatic synergy that abolishes chromosome segregation. This suggests that these two activities jointly coordinate mitotic progression. Accordingly, recent evidence suggests that Aurora A and B work together in both spindle assembly in metaphase and disassembly in anaphase. Here, we provide an outlook on these shared functions of the Auroras, discuss the evolution of this family of mitotic kinases and speculate why Aurora kinase activity may be required at both ends of the spindle microtubules.
- A Genomic Signature and the Identification of New Sporulation GenesPublication . Abecasis, A. B.; Serrano, M.; Alves, R.; Quintais, L.; Pereira-Leal, J. B.; Henriques, A. O.Bacterial endospores are the most resistant cell type known to humans, as they are able to withstand extremes of temperature, pressure, chemical injury, and time. They are also of interest because the endospore is the infective particle in a variety of human and livestock diseases. Endosporulation is characterized by the morphogenesis of an endospore within a mother cell. Based on the genes known to be involved in endosporulation in the model organism Bacillus subtilis, a conserved core of about 100 genes was derived, representing the minimal machinery for endosporulation. The core was used to define a genomic signature of about 50 genes that are able to distinguish endospore-forming organisms, based on complete genome sequences, and we show this 50-gene signature is robust against phylogenetic proximity and other artifacts. This signature includes previously uncharacterized genes that we can now show are important for sporulation in B. subtilis and/or are under developmental control, thus further validating this genomic signature. We also predict that a series of polyextremophylic organisms, as well as several gut bacteria, are able to form endospores, and we identified 3 new loci essential for sporulation in B. subtilis: ytaF, ylmC, and ylzA. In all, the results support the view that endosporulation likely evolved once, at the base of the Firmicutes phylum, and is unrelated to other bacterial cell differentiation programs and that this involved the evolution of new genes and functions, as well as the cooption of ancestral, housekeeping functions.
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