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Distributional shifts in species ranges provide critical evidence of ecological responses to climate change. Assessments of climate-driven changes typically focus on broad-scale range shifts (e.g. poleward or upward), with ecological consequences at regional and local scales commonly overlooked. While these changes are informative for species presenting continuous geographic ranges, many species have discontinuous distributions-both natural (e.g. mountain or coastal species) or human-induced (e.g. species inhabiting fragmented landscapes)-where within-range changes can be significant. Here, we use an ecosystem engineer species (Sabellaria alveolata) with a naturally fragmented distribution as a case study to assess climate-driven changes in within-range occupancy across its entire global distribution. To this end, we applied landscape ecology metrics to outputs from species distribution modelling (SDM) in a novel unified framework. SDM predicted a 27.5% overall increase in the area of potentially suitable habitat under RCP 4.5 by 2050, which taken in isolation would have led to the classification of the species as a climate change winner. SDM further revealed that the latitudinal range is predicted to shrink because of decreased habitat suitability in the equatorward part of the range, not compensated by a poleward expansion. The use of landscape ecology metrics provided additional insights by identifying regions that are predicted to become increasingly fragmented in the future, potentially increasing extirpation risk by jeopardising metapopulation dynamics. This increased range fragmentation could have dramatic consequences for ecosystem structure and functioning. Importantly, the proposed framework-which brings together SDM and landscape metrics-can be widely used to study currently overlooked climate-driven changes in species internal range structure, without requiring detailed empirical knowledge of the modelled species. This approach represents an important advancement beyond predictive envelope approaches and could reveal itself as paramount for managers whose spatial scale of action usually ranges from local to regional.

Microbial communities are a siege of complex metabolic interactions such as cooperation and competition for resources. Methods to control such interactions could lead to major advances in our ability to engineer microbial consortia for bioproduction and synthetic biology applications. Here, we used optogenetics to control invertase production in yeast, thereby creating landscapes of cooperator and cheater cells. Yeast cells behave as cooperators (i.e.,transform sucrose into glucose, a public “good”) upon blue light illumination or cheaters (i.e.,consume glucose produced by cooperators to grow) in the dark. We show that cooperators benefit best from the hexoses they produce when their domain size is constrained between two cut-off length-scales. From an engineering point of view, the system behaves as a band pass filter. The lower limit is the trace of cheaters’ competition for hexoses, while the upper limit is defined by cooperators’ competition for sucrose. Hence, cooperation mostly occurs at the frontiers with cheater cells, which not only compete for hexoses but also cooperate passively by letting sucrose reach cooperators. We anticipate that this optogenetic method could be applied to shape metabolic interactions in a variety of microbial ecosystems.

Natural environments of living organisms are often dynamic and multifactorial, with multiple parameters fluctuating over time. To better understand how cells respond to dynamically interacting factors, we quantified the effects of dual fluctuations of osmotic stress and glucose deprivation on yeast cells using microfluidics and time-lapse microscopy. Strikingly, we observed that cell proliferation, survival and signaling depend on the phasing of the two periodic stresses. Cells divided faster, survived longer and showed decreased transcriptional response when fluctuations of hyperosmotic stress and glucose deprivation occurred in phase than when the two stresses occurred alternatively. We also found that mutants with impaired osmotic stress response were better adapted to alternating stresses than wild-type cells, showing that genetic mechanisms of adaptation to a persistent stress factor can be detrimental under dynamically interacting conditions. Taken together, we demonstrate that glucose availability regulates yeast responses to dynamic osmotic stress. We anticipate that our approach can be extended to other stress responsive pathways to further elucidate the key role of metabolic fluctuations in the dynamics of cellular responses to stress.

Optogenetics arises as a valuable tool to precisely control genetic circuits in microbial cell factories. Light control holds the promise of optimizing bioproduction methods and maximize yields, but its implementation at different steps of the strain development process and at different culture scales remains challenging. In this study, we aim to control beta-carotene bioproduction using optogenetics inSaccharomyces cerevisiaeand investigate how its performance translates across culture scales. We built four lab-scale illumination devices, each handling different culture volumes, and each having specific illumination characteristics and cultivating conditions. We evaluated optogenetic activation and beta-carotene production across devices and optimized them both independently. Then, we combined optogenetic induction and beta-carotene production to make a light-inducible beta-carotene producer strain. This was achieved by placing the transcription of the bifunctional lycopene cyclase / phytoene synthase CrtYB under the control of the pC120 optogenetic promoter regulated by the EL222-VP16 light-activated transcription factor, while other carotenogenic enzymes (CrtI, CrtE, tHMG) were expressed constitutively. We show that illumination, culture volume and shaking impact differently optogenetic activation and beta-carotene production across devices. This enabled us to determine the best culture conditions to maximize light-induced beta-carotene production in each of the devices, reaching a content of up to 880 μg/gCDW. Our study exemplifies the stakes of scaling up optogenetics in devices of different lab scales and sheds light on the interplays and potential conflicts between optogenetic control and metabolic pathway efficiency. As a general principle, we propose that it is important to first optimize both components of the system independently, before combining them into optogenetic producing strains to avoid extensive troubleshooting. We anticipate that our results can help designing both strains and devices that could eventually lead to larger scale systems in an effort to bring optogenetics to the industrial scale.

Timelapse fluorescence microscopy imaging is routinely used in quantitative cell biology. However, microscopes could become much more powerful investigation systems if they were endowed with simple unsupervised decision-making algorithms to transform them into fully responsive and automated measurement devices. Here, we report CyberSco.Py, Python software for advanced automated timelapse experiments. We provide proof-of-principle of a user-friendly framework that increases the tunability and flexibility when setting up and running fluorescence timelapse microscopy experiments. Importantly, CyberSco.Py combines real-time image analysis with automation capability, which allows users to create conditional, event-based experiments in which the imaging acquisition parameters and the status of various devices can be changed automatically based on the image analysis. We exemplify the relevance of CyberSco.Py to cell biology using several use case experiments with budding yeast. We anticipate that CyberSco.Py could be used to address the growing need for smart microscopy systems to implement more informative quantitative cell biology experiments.

Introduction Parmi les differents mecanismes et influences auxquels on attribue un lien avec la douleur chronique, le modele d’evitement de la douleur est considere comme un mecanisme important du developpement de la douleur chronique. Alors que les croyances des patients en ce qui concerne le mal de dos ont ete, et sont toujours etudiees et decrites, nous n’avons que peu d’informations sur les croyances des professionnels de la sante et de leurs possibles repercussions sur les patients. Il y a de bonnes raisons d’etudier ces croyances et attitudes. D’abord, il convient de determiner si les cliniciens se conforment aux recommandations de bonne pratique concernant le traitement du mal de dos chronique emises au terme de differentes recherches. Il convient ensuite de verifier si elles influencent le traitement, la communication et les informations fournies aux patients et si cela joue un role dans la persistance de la douleur. Methode Une etude de la documentation disponible a ete effectuee dans le cadre du diplome de Master es sciences en physiotherapie a l’universite de Brighton (GB). Les bases de donnees Pubmed, Cochrane, Google scholar, MeSH et Pedro ont permis de trouver 15 etudes ; 8 remplissaient les criteres d’inclusion. Resultats Certains therapeutes et medecins semblent encore penser de facon biomedicale, ce qui ne correspond ni aux resultats de la recherche ni aux recommandations de bonne pratique. Il est frappant de constater que certains professionnels de la sante estiment encore que les patients atteints de mal de dos chronique devraient eviter les mouvements douloureux et que le conge maladie constitue une intervention appropriee. De plus, les patients atteints de mal de dos chronique semblent plus susceptibles d’etre des patients « difficiles » du fait de leurs problemes bio-psycho-sociaux complexes et sont laisses a eux-memes plus rapidement que d’autres patients. Conclusions Il est probable que les professionnels de la sante preferent suivre une approche biomedicale parce que cela correspond a leur type de formation. Reconnaitre et traiter les problemes psychologiques peut inquieter les professionnels de la sante car cela depasse leurs connaissances. Il reste a relever aussi qu’une grande partie des professionnels de la sante ne fournissent pas d’informations claires au patient, que ce soit au sujet de la douleur, de l’activite ou du travail. Considerant le fait que de nombreux professionnels de la sante suivent une approche biomedicale face au mal de dos chronique, il ne semble pas surprenant que les patients aient souvent une mauvaise conception de ce probleme. Signification En conclusion, les recherches mettent en evidence le fait que les croyances et attitudes de nombreux professionnels de la sante peuvent avoir un impact sur le traitement et l’education du patient, et par consequent sur le resultat. Les professionnels de la sante devraient donc etre conscients de leur influence et de leur responsabilite vis-a-vis des patients. Ceux-ci ont le droit d’obtenir le meilleur traitement possible, ce qui implique des informations claires et non ambigues sur le mal de dos chronique et sa gestion.

International audience; Peptide nucleic acids (PNAs) are very attractive antisense and antigene agents, but these molecules are not passively taken into cells. Here, using a functional cell assay and fluorescent-based methods, we investigated cell uptake and antisense activity of a tridecamer PNA that targets the HIV-1 polypurine tract sequence delivered using the arginine-rich (R/W)9 peptide (RRWWRRWRR). At micromolar concentrations, without use of any transfection agents, almost 80% inhibition of the target gene expression was obtained with the conjugate in the presence of the endosomolytic agent chloroquine. We show that chloroquine not only induced escape from endosomes but also enhanced the cellular uptake of the conjugate. Mechanistic studies revealed that (R/W)9-PNA conjugates were internalized via pinocytosis. Replacement of arginines with lysines reduced the uptake of the conjugate by six-fold, resulting in the abolition of intracellular target inhibition. Our results show that the arginines play a crucial role in the conjugate uptake and antisense activity. To determine whether specificity of the interactions of arginines with cell surface proteoglycans result in the internalization, we used flow cytometry to examine uptake of arginine-and lysine-rich conjugates in wild-type CHO-K1 and proteoglycan-deficient A745 cells. The uptake of both conjugates was decreased by four fold in CHO-745 cells; therefore proteoglycans promote internalization of cationic peptides, irrespective of the chemical nature of their positive charges. Our results show that arginine-rich cell-penetrating peptides, especially (R/W)9, are a promising tool for PNA internalization.

Oligonucleotides and their analogues, such as peptide nucleic acids (PNAs), can be used in chemical strategies to artificially control gene expression. Inefficient cellular uptake and inappropriate cellular localization still remain obstacles in biological applications, however, especially for PNAs. Here we demonstrate that conjugation of PNAs to flavin resulted in efficient internalization into cells through an endocytic pathway. The flavin-PNAs exhibited antisense activity in the sub-micromolar range, in the context of a treatment facilitating endosomal escape. Increased endosomal release of flavin conjugates into the cytoplasm and/or nucleus was shown by chloroquine treatment and also--when the flavin-PNA was conjugated to rhodamine, a mild photosensitizer--upon light irradiation. In conclusion, an isoalloxazine moiety can be used as a carrier and attached to a cargo biomolecule, here a PNA, for internalization and functional cytoplasmic/nuclear delivery. Our findings could be useful for further design of PNAs and other oligonucleotide analogues as potent antisense agents.

The insulin-like growth factor 1 receptor (IGF-1R) is involved in transformation, survival, mitogenesis and differentiation. It is overexpressed in many tumors and a validated target for anticancer therapy. In cell-free systems, polypyrimidic peptide nucleic acids (PNAs) can form triplex-like structures with messenger RNAs and halt the ribosomal machinery during the translation elongation. A 17-mer PNA that formed a PNA(2):mRNA complex with a purine-rich sequence located in the coding region of IGF-1R mRNA induced the synthesis of a truncated IGF-1R in vitro. This PNA down-regulated expression of the receptor by 70-80% in prostate cancer cells without affecting insulin receptor expression that exhibits high homology with IGF-1R. Inhibition occurs at the translational level, since the IGF-1R mRNA level measured by quantitative RT-PCR was not affected by PNA treatment. In addition, IGF-1R knockdown by PNA led to an attenuation of phosphorylation of downstream signaling pathways, PI3K/AKT and MAPK, involved in survival and mitogenesis and also to a decrease in cell transformation. Of the steric blockers tested, which included phosphorodiamidate morpholino oligomers and locked nucleic acids, PNA was unique in its ability to form triplex structures with mRNA and to arrest translation elongation.