Drought tolerance conferred to sugarcane by association with Gluconacetobacter diazotrophicus: a transcriptomic view of hormone pathways

Sugarcane interacts with particular types of beneficial nitrogen-fixing bacteria that provide fixed-nitrogen and plant growth hormones to host plants, promoting an increase in plant biomass. Other benefits, as enhanced tolerance to abiotic stresses have been reported to some diazotrophs. Here we aim to study the effects of the association between the diazotroph Gluconacetobacter diazotrophicus PAL5 and sugarcane cv. SP70-1143 during water depletion by characterizing differential transcriptome profiles of sugarcane. RNA-seq libraries were generated from roots and shoots of sugarcane plants free of endophytes that were inoculated with G. diazotrophicus and subjected to water depletion for 3 days. A sugarcane reference transcriptome was constructed and used for the identification of differentially expressed transcripts. The differential profile of non-inoculated SP70-1143 suggests that it responds to water deficit stress by the activation of drought-responsive markers and hormone pathways, as ABA and Ethylene. qRT-PCR revealed that root samples had higher levels of G. diazotrophicus 3 days after water deficit, compared to roots of inoculated plants watered normally. With prolonged drought only inoculated plants survived, indicating that SP70-1143 plants colonized with G. diazotrophicus become more tolerant to drought stress than non-inoculated plants. Strengthening this hypothesis, several gene expression responses to drought were inactivated or regulated in an opposite manner, especially in roots, when plants were colonized by the bacteria. The data suggests that colonized roots would not be suffering from stress in the same way as non-inoculated plants. On the other hand, shoots specifically activate ABA-dependent signaling genes, which could act as key elements in the drought resistance conferred by G. diazotrophicus to SP70-1143. This work reports for the first time the involvement of G. diazotrophicus in the promotion of drought-tolerance to sugarcane cv. SP70-1143, and it describes the initial molecular events that may trigger the increased drought tolerance in the host plant

Relative roles of genetic and epigenetic variation on the ecology and evolution of mangrove killifishes (Kryptolebias spp.)

The field of ecological epigenetics aims to understand the implications of epigenetic modifications in adaptation, inheritance and ultimately, evolution. Many questions remain open within ecological epigenetics, in particular, how epigenetic variation is influenced by genetic background, the extent of environmentally-induced epigenetic variants, as well as its degree of heritability. This thesis used the unique diversity of mating systems present in the killifish genus Kryptolebias to investigate how genetic and environmental variation shape epigenetic variation in animals. Genetic and epigenetic structure was investigated in natural populations of K. hermaphroditus in northeast Brazil, with the species being confirmed as the second example of mixed-mating system in vertebrates. Cytosine methylation was largely influenced by genetic background. However, within-populations, when individuals were more genetically similar, DNA methylation was mostly affected by parasites. Kryptolebias ocellatus, here confirmed as an outcrossing-only androdioecious species, showed deep genetic structure in southeast Brazil. Hybridisation between K. ocellatus and the predominantly selfing K. hermaphroditus was uncovered, representing the first example of hybridisation between species with different mating systems in vertebrates. Hybrids had intermediate patterns of cytosine methylation relatively to the parental species, with important biological processes being potentially misregulated. Environmental enrichment was shown to affect brain cytosine methylation patterns in two inbred strains of K. marmoratus, however genetic background had a stronger effect than environmental variation. Commonly-affected epialleles between genotypes predominantly showed a genotype-by-environment reaction norm, suggesting that exclusively environmentally-induced epialleles may be rare. Intergenerationally, parental activity affected offspring activity, and a limited number of putative intergenerational epialleles were identified. This is the first example of behavioural parental effects induced by environmental enrichment in fish. These findings show that genetic background has a prominent effect and must be take into account when evaluating the evolutionary potential of cytosine methylation variation. In addition, inheritance of environmentally-induced cytosine methylation epialleles may be limited, with other epigenetic mechanisms, such as microRNAs, being more likely to escape epigenetic reprogramming and transmit epigenetically-induced parental effects

Sperm Proteome Maturation in the Mouse Epididymis

abstract: In mammals, transit through the epididymis, which involves the acquisition, loss and modification of proteins, is required to confer motility and fertilization competency to sperm. The overall dynamics of maturation is poorly understood, and a systems level understanding of the complex maturation process will provide valuable new information about changes occurring during epididymal transport. We report the proteomes of sperm collected from the caput, corpus and cauda segments of the mouse epididymis, identifying 1536, 1720 and 1234 proteins respectively. This study identified 765 proteins that are present in sperm obtained from all three segments. We identified 1766 proteins that are potentially added (732) or removed (1034) from sperm during epididymal transit. Phenotypic analyses of the caput, corpus and cauda sperm proteomes identified 60 proteins that have known sperm phenotypes when mutated, or absent from sperm. Our analysis indicates that as much as one-third of proteins with known sperm phenotypes are added to sperm during epididymal transit. GO analyses revealed that cauda sperm are enriched for specific functions including sperm-egg recognition and motility, consistent with the observation that sperm acquire motility and fertilization competency during transit through the epididymis. In addition, GO analyses revealed that the immunity protein profile of sperm changes during sperm maturation. Finally, we identified components of the 26S proteasome, the immunoproteasome, and a proteasome activator in mature sperm.The article is published at http://journals.plos.org/plosone/article?id=10.1371/journal.pone.014065

AGI for Agriculture

Artificial General Intelligence (AGI) is poised to revolutionize a variety of sectors, including healthcare, finance, transportation, and education. Within healthcare, AGI is being utilized to analyze clinical medical notes, recognize patterns in patient data, and aid in patient management. Agriculture is another critical sector that impacts the lives of individuals worldwide. It serves as a foundation for providing food, fiber, and fuel, yet faces several challenges, such as climate change, soil degradation, water scarcity, and food security. AGI has the potential to tackle these issues by enhancing crop yields, reducing waste, and promoting sustainable farming practices. It can also help farmers make informed decisions by leveraging real-time data, leading to more efficient and effective farm management. This paper delves into the potential future applications of AGI in agriculture, such as agriculture image processing, natural language processing (NLP), robotics, knowledge graphs, and infrastructure, and their impact on precision livestock and precision crops. By leveraging the power of AGI, these emerging technologies can provide farmers with actionable insights, allowing for optimized decision-making and increased productivity. The transformative potential of AGI in agriculture is vast, and this paper aims to highlight its potential to revolutionize the industry

CERNAS – Current Evolution and Research Novelty in Agricultural Sustainability

This book addresses original studies and reviews focused on the current evolution and research novelty in agricultural sustainability. New developments are discussed on issues related with quality of soil, natural fertilizers or the sustainable use of land and water. Also crop protection techniques are pivotal for the sustainable food production under the challenges of the Sustainable Development Goals of the United Nations, allied to innovative weed control methodologies, as a way to reduce the utilization of pesticides. The role of precision and smart agriculture is becoming more pertinent as the communication technologies improve at a high rate. Waste management, reuse of agro industrial residues, extension of shelf life and use of new technologies are ways to reduce food waste, all contributing to a higher sustainability of the food supply chains, leading to a more rational use of natural resources. The unquestionable role of bees as pollinators and contributors for biodiversity is subjacent to the work of characterization of beekeeping activities, which in turn contribute, together with the valorization of endemic varieties of plant foods, for the development of local communities. Finally, the short circuits and local food markets have a decisive role in the preservation and enhancement of rural economies.info:eu-repo/semantics/publishedVersio

CERNAS: Current Evolution and Research Novelty in Agricultural Sustainability

Climate changes pose overwhelming impacts on primary production and, consequently, on agricultural and animal farming. Additionally, at present, agriculture still depends strongly on fossil fuels both for energy and production factors ,such as synthetized inorganic fertilizers and harmful chemicals such as pesticides. The need to feed the growing world population poses many challenges. The need to reduce environmental impacts to a minimum, maintain healthy ecosystems, and improve soil microbiota are central to ensuring a promising future for coming generations. Livestock production under cover crop systems helps to alleviate compaction so that oxygen and water can sufficiently flow in the soil, add organic matter, and help hold soil in place, reducing crusting and protecting against erosion. The use of organic plant production practices allied to the control of substances used in agriculture also decisively contributes to alleviating the pressure on ecosystems. Some of the goals of this new decade are to use enhanced sustainable production methodologies to improve the input/output ratios of primary production, reduce environmental impacts, and rely on new innovative technologies. This reprint addresses original studies and reviews focused on the current evolution and research novelty in agricultural sustainability. New developments are discussed on issues related to quality of soil, natural fertilizers, or the sustainable use of land and water. Also, crop protection techniques are pivotal for sustainable food production under the challenges of the Sustainable Development Goals of the United Nations, allied to innovative weed control methodologies as a way to reduce the utilization of pesticides. The role of precision and smart agriculture is becoming more pertinent as communication technologies improve at a rapid rate. Waste management, reuse of agro-industrial residues, extension of shelf life, and use of new technologies are ways to reduce food waste, all contributing to higher sustainability in food supply chains, leading to a more rational use of natural resources. The unquestionable role of bees as pollinators and contributors to biodiversity is adjacent to characterizing beekeeping activities, which in turn contributes, together with the valorization of endemic varieties of plant foods, to the development of local communities. Finally, the short circuits and local food markets have a decisive role in the preservation and enhancement of rural economies.info:eu-repo/semantics/publishedVersio

A Survey of the First 20 Years of Research on Semantic Web and Linked Data

International audienceThis paper is a survey of the research topics in the field of Semantic Web, Linked Data and Web of Data. This study looks at the contributions of this research community over its first twenty years of existence. Compiling several bibliographical sources and bibliometric indicators , we identify the main research trends and we reference some of their major publications to provide an overview of that initial period. We conclude with some perspectives for the future research challenges.Cet article est une étude des sujets de recherche dans le domaine du Web sémantique, des données liées et du Web des données. Cette étude se penche sur les contributions de cette communauté de recherche au cours de ses vingt premières années d'existence. En compilant plusieurs sources bibliographiques et indicateurs bibliométriques, nous identifions les principales tendances de la recherche et nous référençons certaines de leurs publications majeures pour donner un aperçu de cette période initiale. Nous concluons avec une discussion sur les tendances et perspectives de recherche

RNA-SEQ applied to the peacock blenny Salaria pavo: unveiling the gene networks and signalling pathways behind phenotypic plasticity in a littoral fish

Phenotypic plasticity is the ability of an individual genome to produce different phenotypes depending on environmental cues. These plastic responses rely on diverse genomic mechanisms and allow an organism to maximize its fitness in a variety of social and physical settings. The development of next-generation sequencing (NGS) technologies, especially RNA Sequencing (RNA-Seq), has made it possible to investigate the distinct patterns of gene expression known to be underlying plastic phenotypes in species with ecological interest. In teleost fishes, changes in phenotypes is often observed during the reproductive season, with shifts and adjustments in dominance status that can lead to the co-existence of multiple reproductive morphs within the same population. One such example is the peacock blenny Salaria pavo (Risso, 1810), a species where the intensity of mating competition varies among populations due to nest-site availability, such that two different levels of plasticity arise: 1) intraspecific variation in reproductive behaviour for males that can follow either of two developmental pathways, grow directly into nest-holder males, or behave first as female mimics to sneak fertilizations (sneaker males) and later transition into nest-holder males, and 2) inter-population variation in courting roles of females and nest-holder males. This system provides the ideal basis to apply RNA-Seq methods to study plasticity since differences in reproductive traits within and among populations can reveal which genetic and genomic mechanisms underpin the observed variation in behavioural response to changes in the social environment. However, the genomic information available for this species was scarce, and hence multiple sequencing techniques were used and the methodologies applied optimized throughout the work. In this thesis, we start by first obtaining a de novo transcriptome assembly to develop the first genetic markers for this species (Chapter 2). These microsatellites were used to elucidate the reproductive success (i.e. consisting of mating success and fertilization success) of male ARTs, which can be used as a proxy of Darwinian fitness (Chapter 3). In this study, we detected a fertilization success for nestholder males of 95%, and showed a stronger influence of the social environment rather than morphological variables in the proportion of lost fertilizations by nest-holder males of this species. Taking advantage of the developed transcriptome, we used highthroughput sequencing to obtain expression profiles for male morphs (i.e. intraspecific variation) and females in this species, and focus on the role of differential gene expression in the evolution of sequential alternative reproductive tactics (ARTs) that involve the expression of both male and female traits (Chapter 4). Additionally, we show how the distinct behavioural repertoires are facilitated by distinct neurogenomic states, which discriminate not only sex but also male morphs. Lastly, using two different target tissues, gonads and forebrain, we focus on the genomic regulation of sex roles in courtship behaviour between females and males from two populations under different selective regimes (inter-population variation), the Portuguese coastal population with reversed sex roles and the rocky Italian population with ‘conventional’ sex roles (Chapter 5). Here we demonstrate that variation in gene expression at the brain level segregates individuals by population rather than by sex, indicating that plasticity in behaviour across populations drives variation in neurogenomic expression. On the other hand, at the gonad level, variation in gene expression segregates individuals by sex and then by population, indicating that sexual selection is also acting at the intrasexual level, particularly in nestholder males by paralleling differences in gonadal investment. However, the genomic mechanisms underlying courtship behaviour were not fully elucidated, and more studies are necessary.A plasticidade fenotípica consiste na capacidade de o mesmo genoma produzir diferentes fenótipos comportamentais dependendo das pistas ambientais recebidas. Estas respostas plásticas dependem de diversos mecanismos genómicos e permitem que o indivíduo maximize a sua fitness (aptidão) numa variedade de ambientes ecológicos. O desenvolvimento verificado nas tecnologias de sequenciação de alto desempenho ao longo da última década, globalmente denominadas de “Next Generation Sequencing” (NGS), permitiu o estabelecimento de métodos de análise e ferramentas genómicas que podem ser aplicadas em todos os sistemas ecológicos de interesse em biologia, sem a existência prévia de um genoma curado. Nomeadamente a tecnologia de sequenciação de ARN, conhecida globalmente como RNA-Seq, tornou possível a investigação dos perfis de expressão génica que se sabe serem determinantes na emergência de fenótipos plásticos, e consequentemente permitem determinar fenótipos em estados distintos de expressão genómica. Em peixes teleósteos, é possível observar com frequência modificações no fenótipo comportamental durante o período de reprodução, como por exemplo alterações e ajustes no estatuto de dominância que podem levar à coexistência de indivíduos que apresentam diferentes táticas de reprodução dentro da mesma população. Um desses exemplos é o peixe marachomba-pavão Salaria pavo (Risso, 1810), onde a intensidade na competição intra e intersexual varia entre populações sendo modulada pela disponibilidade de locais de nidificação, de forma a que dois níveis diferentes de plasticidade surgem: 1) variação intraespecífica no comportamento reprodutivo em machos que podem seguir uma de duas vias de desenvolvimento, investirem no seu crescimento e tornarem-se machos nidificantes na sua primeira época de reprodução, ou primeiro seguir uma tática de macho parasita onde investem em fertilizações furtivas, sendo que mais tarde no seu desenvolvimento fazem a transição para macho nidificante, e 2) variação interpopulacional nos papeis de corte de fêmeas e machos nidificantes. Os machos parasitas, conhecidos nesta espécie como “sneakers”, possuem uma particularidade que os tornam singulares, para além de imitarem a morfologia das fêmeas também conseguem imitar o seu comportamento de corte direcionado ao macho nidificante, o que lhes permite aproximarem-se discretamente dos ninhos dos machos e fertilizar parte dos ovos que as fêmeas depositam. Este sistema constitui a base ideal para aplicar métodos de RNA-Seq e estudar esta plasticidade fenotípica, uma vez que diferenças nas características reprodutivas dentro e entre populações podem revelar quais os mecanismos genéticos e genómicos subjacentes à variação observada em resposta a mudanças no ambiente ecológico. No entanto, a informação genómica disponível nesta espécie é reduzida e, por isso diferentes técnicas de sequenciação, assim como diferentes métodos de análise foram usados e otimizados ao longo deste trabalho. A presente tese é constituída por quatro trabalhos, sendo que no primeiro estudo se começa pela sequenciação de uma biblioteca de ARN proveniente de uma mistura de múltiplos indivíduos e de tecidos, de forma a captar a diversidade genética e desenvolver os primeiros marcadores genéticos nesta espécie (Capítulo 2). Com base nestes marcadores, microssatélites polimórficos, foi possível genotipar uma fração dos indivíduos da população existente na Ilha da Culatra (Ria Formosa, Portugal) bem como os ovos retirados de ninhos alvo, de forma a fazer análises de paternidade (Capítulo 3). Neste estudo, foi possível estimar o sucesso de fertilização de ovos de cada uma das táticas alternativas de reprodução, e usá-la como medida representativa de fitness de cada tática alternativa de reprodução nesta espécie. Os resultados indicam um sucesso de fertilização para os machos nidificantes de 95%, e mostramos que existe uma maior influência do ambiente social do que de variáveis morfológicas na proporção de fertilizações não obtidas pelos machos nidificantes, quando comparado com estudos anteriores. Usando o transcriptoma obtido no primeiro trabalho, avançámos com a caraterização genómica de cada um dos fenótipos presentas na população da ilha da Culatra, fêmeas, machos nidificantes, machos sneakers e machos de transição (machos que apenas investem no seu crescimento, não se reproduzindo, e consequente transição de sneaker para macho nidificante) (Capítulo 4). Para tal, foi sequenciado em profundidade o transcriptoma de cérebro de cada um deste fenótipos, e os perfis de expressão obtidos para machos e fêmeas desta espécie, onde o foco do estudo se centrava no papel da expressão génica diferencial na evolução de táticas reprodutivas alternativas sequenciais que envolvem a expressão de ambos os traços masculinos e femininos. Os resultados obtidos, mostram como repertórios comportamentais distintos são facilitados por estados neurogenómicos distintos, que discriminam não apenas o sexo, mas também as táticas alternativas de reprodução. Por fim, utilizando dois tecidos-alvo, gónadas e prosencéfalo, focámo-nos na regulação genómica dos papeis sexuais no comportamento de corte entre fêmeas e machos nidificantes de duas populações sob diferentes regimes seletivos, a população costeira portuguesa com papeis sexuais invertidos e a população rochosa italiana, com papeis sexuais ‘convencionais’ (Capítulo 5). Os resultados obtidos mostram que ao nível do cérebro, a variação na expressão génica segrega os indivíduos por população e não por sexo, indicando que a plasticidade no comportamento entre as populações induz uma maior variação na expressão neurogenómica. Por outro lado, ao nível das gónadas, a variação na expressão génica segrega os indivíduos por sexo e também por população, indicando que a seleção sexual está a atuar ao nível intrasexual, particularmente nos machos nidificantes, indo de encontro a diferenças detetadas entre populações no investimento alocado às gónadas. No entanto, os mecanismos genómicos subjacentes ao comportamento de corte não foram totalmente elucidados, e mais estudos são necessários.The work presented here was developed at Instituto Gulbenkian de Ciência (IGC) in Oeiras, with the support of both ISPA – Instituto Universitário in Lisbon, for the maintenance of live fish, and Centro de Ciências do Mar (CCMAR) at Universidade do Algarve, for logistics and support during fieldwork in Ria Formosa

Abordagem à regulação da mobilidade do espermatozoide através da caracterização e modulação da vis de sinalização GSK/PPP1R2/PPP1

Doutoramento em BiologiaSperm motility acquisition and maintenance is a fundamental process for oocyte fertilization and consequently conception. The signaling events underling sperm motility acquisition have been studied for decades. However, many questions are still unanswered. Also, the limited options currently available for male contraception (condom, vasectomy and withdrawal) reflect the necessity of a new group of male contraceptives based on sperm motility modulation. GSK3/PPP1R2/PPP1 signaling pathway is involved in sperm motility acquisition during epididymis transit. The main goal for this work was to deepen the knowledge on the signaling events involved in human sperm motility by focusing on the characterization and modulation of the signaling pathway GSK3/PPP1R2/PPP1. We first designed, synthetized and characterized a disruptive bioportide based on cell penetrating peptide technology. In vitro studies revealed that the disruptive bioportide interferes with PPP1R2/PPP1CC2 interaction and restores PPP1CC2 activity. We also demonstrated that when exposed to the disruptive bioportide, sperm motility is significantly reduced. Aiming to identify sperm protein-protein interactions suitable for pharmacological intervention, we turn our attention to GSK3, a modulator of PPP1R2/PPP1CC2 interactions in sperm. We provide for the first time GSK3 human testis and sperm interactomes. We reported an isoforms specific role for GSK3 in human sperm motility and an in silico analysis revealed GSK3 and GSK3 interactions involved in sperm motility and potential targets for pharmacological intervention. In conclusion, we demonstrated that it is possible to target protein-protein interactions and modulate sperm complexes involved in motility using bioportides. Moreover, we identified new potential protein interactions involved in sperm motility and showed that the development of new type of male contraceptive based on inhibiting sperm motility is now achievable.A aquisição e manutenção da mobilidade do espermatozoide é fundamental para a fertilização do oócito e consequentemente conceção. Durante décadas, as vias de sinalização necessárias à aquisição de mobilidade por parte do espermatozoide foram alvo de intensos estudos. Contudo, este processo ainda não é inteiramente conhecido. Ademais, as limitadas opções disponíveis para contraceção masculina (preservativo, vasectomia e coito interrompido) refletem a necessidade de desenvolver um contracetivo masculino baseado na modulação da mobilidade do espermatozoide. A via de sinalização GSK3/PPP1R2/PPP1 está envolvida na aquisição de mobilidade do espermatozoide ao longo do transito do epidídimo. O objetivo principal deste trabalho é enriquecer o conhecimento dos eventos celulares necessários na mobilidade do espermatozoide através da caracterização e modulação da via de sinalização GSK3/PPP1R2/PPP1 em espermatozoides humanos. Desenhámos, sintetizámos e caracterizámos um bioportide que quebra interações proteicas baseado em tecnologia de cell penetrating peptides. Estudos in vitro revelaram que o bioportide de ruptura interfere com a interação PPP1R2/PPP1CC2 e é capaz de restabelecer a atividade da PPP1CC2. Também demonstramos que o bioportide reduz significativamente a mobilidade do espermatozoide. Com o objetivo de identificar interacções proteína-proteína adequadas à intervenção farmacológica, focámos a nossa atenção na proteína GSK3, um modulador da interação PPP1R2/PPP1CC2 em espermatozoides. Descrevemos pela primeira vez o interactoma da GSK3 no testículo e espermatozoide humanos e reportamos um papel específico da isoforma GSK3 na mobilidade do espermatozoide. Uma análise in silico revelou interatores da GSK3 e GSK3 que estão envolvidos na mobilidade do espermatozoide e potencialmente poderão ser alvos de intervenção farmacológica para um novo contraceptivo masculino. Em conclusão, demonstramos que é possível provocar a quebra de interações proteína-proteína e modular a mobilidade do espermatozoide usando de bioportides. Também identificamos potenciais novas interações proteicas envolvidas na mobilidade do espermatozoide. Finalmente, mostramos que é possível idealizar um novo tipo de contracepção masculina baseado na inibição da mobilidade do espermatozoide