Precursor uptake assays and metabolic analyses in isolated tomato fruit chromoplasts

Background Carotenoids are the most widespread group of pigments found in nature. In addition to their role in the physiology of the plant, carotenoids also have nutritional relevance as their incorporation in the human diet provides health benefits. In non-photosynthetic tissues, carotenoids are synthesized and stored in specialized plastids called chromoplasts. At present very little is known about the origin of the metabolic precursors and cofactors required to sustain the high rate of carotenoid biosynthesis in these plastids. Recent proteomic data have revealed a number of biochemical and metabolic processes potentially operating in fruit chromoplasts. However, considering that chloroplast to chromoplast differentiation is a very rapid process during fruit ripening, there is the possibility that some of the proteins identified in the proteomic analysis could represent remnants no longer having a functional role in chromoplasts. Therefore, experimental validation is necessary to prove whether these predicted processes are actually operative in chromoplasts. Results A method has been established for high-yield purification of tomato fruit chromoplasts suitable for metabolic studies. Radiolabeled precursors were efficiently incorporated and further metabolized in isolated chromoplast. Analysis of labeled lipophilic compounds has revealed that lipid biosynthesis is a very efficient process in chromoplasts, while the relatively low incorporation levels found in carotenoids suggest that lipid production may represent a competing pathway for carotenoid biosynthesis. Malate and pyruvate are efficiently converted into acetyl-CoA, in agreement with the active operation of the malic enzyme and the pyruvate dehydrogenase complex in the chromoplast. Our results have also shown that isolated chromoplasts can actively sustain anabolic processes without the exogenous supply of ATP, thus suggesting that these organelles may generate this energetic cofactor in an autonomous way. Conclusions We have set up a method for high yield purification of intact tomato fruit chromoplasts suitable for precursor uptake assays and metabolic analyses. Using targeted radiolabeled precursors we have been able to unravel novel biochemical and metabolic aspects related with carotenoid and lipid biosynthesis in tomato fruit chromoplasts. The reported chromoplast system could represent a valuable platform to address the validation and characterization of functional processes predicted from recent transcriptomic and proteomic data.

Genome-wide DNA polymorphism analyses using VariScan

BACKGROUND: DNA sequence polymorphisms analysis can provide valuable information on the evolutionary forces shaping nucleotide variation, and provides an insight into the functional significance of genomic regions. The recent ongoing genome projects will radically improve our capabilities to detect specific genomic regions shaped by natural selection. Current available methods and software, however, are unsatisfactory for such genome-wide analysis. RESULTS: We have developed methods for the analysis of DNA sequence polymorphisms at the genome-wide scale. These methods, which have been tested on a coalescent-simulated and actual data files from mouse and human, have been implemented in the VariScan software package version 2.0. Additionally, we have also incorporated a graphical-user interface. The main features of this software are: i) exhaustive population-genetic analyses including those based on the coalescent theory; ii) analysis adapted to the shallow data generated by the high-throughput genome projects; iii) use of genome annotations to conduct a comprehensive analyses separately for different functional regions; iv) identification of relevant genomic regions by the sliding-window and wavelet-multiresolution approaches; v) visualization of the results integrated with current genome annotations in commonly available genome browsers. CONCLUSION: VariScan is a powerful and flexible suite of software for the analysis of DNA polymorphisms. The current version implements new algorithms, methods, and capabilities, providing an important tool for an exhaustive exploratory analysis of genome-wide DNA polymorphism data.

L'exploitation des ressources végétales dans les Pyrénées orientales durant l'holocène: analyse anthracologique des structures d'élevage, de four et de charbonnières dans l'Alt Urgell (Chaîne du Cadí) et la Vallée du Madriu (Andorre)

Dans ce travail, nous présentons le résultat des recherches anthracologiques réalisées sur des sites archéologiques de haute montagne dans les Pyrénées orientales. Cette recherche s’insère dans un projet d’archéologie du paysage. Les zones d’étude se situent dans les Pré-Pyrénées sur le versant méridional de la chaîne du Cadí (vallée de la Vansa, Alt Urgell), et dans la vallée du Madriu, située dans les Pyrénées axiales, en Andorre. Le but principal de l’analyse anthracologique a été de connaître et comprendre la dynamique et la relation qui unissent les populations qui ont occupé ces vallées avec le milieu forestier de haute montagne. Les échantillons ont été récupérés sur des structures archéologiques associés à l’exploitation forestière (charbonnières, fours pour la fabrication de résine), au pastoralisme (cabanes et enclos) et à l’exploitation minière-métallurgique (four de grillage), avec une chronologie qui débute au Néolithique Ancien et finit à l’époque moderne et contemporaine (XVIII-XIXe siècles). Les résultats indiquent une variabilité taxonomique pauvre avec la présence majoritaire du Pinus et la présence ponctuelle d’autres espèces arbustives (Ericaceae et Juniperus) et arborées, comme Betula alba ou Abies alba. Différentes formations végétales de haute montagne ont été identifiées, selon leur localisation en versant ensoleillé ou ombragé, ainsi que leur altitude. Les résultats montrent des traces d’exploitation forestière dès le Néolithique Ancien, une intensification des activités à l’Antiquité, même si l’impact anthropique majeur correspond à l’activité de charbonnage d’époque moderne.

Genome-wide DNA polymorphism analyses using VariScan

BACKGROUND: DNA sequence polymorphisms analysis can provide valuable information on the evolutionary forces shaping nucleotide variation, and provides an insight into the functional significance of genomic regions. The recent ongoing genome projects will radically improve our capabilities to detect specific genomic regions shaped by natural selection. Current available methods and software, however, are unsatisfactory for such genome-wide analysis. RESULTS: We have developed methods for the analysis of DNA sequence polymorphisms at the genome-wide scale. These methods, which have been tested on a coalescent-simulated and actual data files from mouse and human, have been implemented in the VariScan software package version 2.0. Additionally, we have also incorporated a graphical-user interface. The main features of this software are: i) exhaustive population-genetic analyses including those based on the coalescent theory; ii) analysis adapted to the shallow data generated by the high-throughput genome projects; iii) use of genome annotations to conduct a comprehensive analyses separately for different functional regions; iv) identification of relevant genomic regions by the sliding-window and wavelet-multiresolution approaches; v) visualization of the results integrated with current genome annotations in commonly available genome browsers. CONCLUSION: VariScan is a powerful and flexible suite of software for the analysis of DNA polymorphisms. The current version implements new algorithms, methods, and capabilities, providing an important tool for an exhaustive exploratory analysis of genome-wide DNA polymorphism data.

Transcriptome analyses of primitively eusocial wasps reveal novel insights into the evolution of sociality and the origin of alternative phenotypes

BACKGROUND: Understanding how alternative phenotypes arise from the same genome is a major challenge in modern biology. Eusociality in insects requires the evolution of two alternative phenotypes - workers, who sacrifice personal reproduction, and queens, who realize that reproduction. Extensive work on honeybees and ants has revealed the molecular basis of derived queen and worker phenotypes in highly eusocial lineages, but we lack equivalent deep-level analyses of wasps and of primitively eusocial species, the latter of which can reveal how phenotypic decoupling first occurs in the early stages of eusocial evolution. RESULTS: We sequenced 20 Gbp of transcriptomes derived from brains of different behavioral castes of the primitively eusocial tropical paper wasp Polistes canadensis. Surprisingly, 75% of the 2,442 genes differentially expressed between phenotypes were novel, having no significant homology with described sequences. Moreover, 90% of these novel genes were significantly upregulated in workers relative to queens. Differential expression of novel genes in the early stages of sociality may be important in facilitating the evolution of worker behavioral complexity in eusocial evolution. We also found surprisingly low correlation in the identity and direction of expression of differentially expressed genes across similar phenotypes in different social lineages, supporting the idea that social evolution in different lineages requires substantial de novo rewiring of molecular pathways. CONCLUSIONS: These genomic resources for aculeate wasps and first transcriptome-wide insights into the origin of castes bring us closer to a more general understanding of eusocial evolution and how phenotypic diversity arises from the same genome.

Trophic structure in a seabird host-parasite food web: insights from stable isotope analyses

Ecological studies on food webs rarely include parasites, partly due to the complexity and dimensionality of host-parasite interaction networks. Multiple co-occurring parasites can show different feeding strategies and thus lead to complex and cryptic trophic relationships, which are often difficult to disentangle by traditional methods. We analyzed stable isotope ratios of C (13C/12C, δ13C) and N (15N/14N, δ15N) of host and ectoparasite tissues to investigate trophic structure in 4 co-occurring ectoparasites: three lice and one flea species, on two closely related and spatially segregated seabird hosts (Calonectris shearwaters). δ13C isotopic signatures confirmed feathers as the main food resource for the three lice species and blood for the flea species. All ectoparasite species showed a significant enrichment in δ15N relatively to the host tissue consumed (discrimination factors ranged from 2 to 5 depending on the species). Isotopic differences were consistent across multiple host-ectoparasite locations, despite of some geographic variability in baseline isotopic levels. Our findings illustrate the influence of both ectoparasite and host trophic ecology in the isotopic structuring of the Calonectris ectoparasite community. This study highlights the potential of stable isotope analyses in disentangling the nature and complexity of trophic relationships in symbiotic systems.

Unique features of odorant-binding proteins of the parasitoid wasp Nasonia vitripennis revealed by genome annotation and comparative analyses

Insects are the most diverse group of animals on the planet, comprising over 90% of all metazoan life forms, and have adapted to a wide diversity of ecosystems in nearly all environments. They have evolved highly sensitive chemical senses that are central to their interaction with their environment and to communication between individuals. Understanding the molecular bases of insect olfaction is therefore of great importance from both a basic and applied perspective. Odorant binding proteins (OBPs) are some of most abundant proteins found in insect olfactory organs, where they are the first component of the olfactory transduction cascade, carrying odorant molecules to the olfactory receptors. We carried out a search for OBPs in the genome of the parasitoid wasp Nasonia vitripennis and identified 90 sequences encoding putative OBPs. This is the largest OBP family so far reported in insects. We report unique features of the N. vitripennis OBPs, including the presence and evolutionary origin of a new subfamily of double-domain OBPs (consisting of two concatenated OBP domains), the loss of conserved cysteine residues and the expression of pseudogenes. This study also demonstrates the extremely dynamic evolution of the insect OBP family: (i) the number of different OBPs can vary greatly between species; (ii) the sequences are highly diverse, sometimes as a result of positive selection pressure with even the canonical cysteines being lost; (iii) new lineage specific domain arrangements can arise, such as the double domain OBP subfamily of wasps and mosquitoes.

Plant uses in different bronze and iron age settlements from the Nuoro province (Sardinia). The results of phytolith analyses from several ceramic fragments and grinding stones.

Ceramic vessels and milling stones are important components of the archaeological record in several Nuraghi from the Pranemuru Plateau (Sardinia). To obtain information on the possible uses of the milling stones and the content vessels is of great interest to understand the economical activities carried out in these sites by these populations. One of the approaches to obtain information on the plant uses was the phytolith analyses of the sediment adhered both to the surface of the milling stones and to the surface of the vessel content. In total we analyzed eleven archaeological samples and two control samples collected from five different Nuraghi in the Pranemuru Plateau (Nuoro Province, Sardinia). The Nuraghi were located in an area of 10 km radius from nuraghe Arrubiu and were chronologically ascribed to the Bronze Age and one site -Pranu Illixi- to the Iron Age.

Health-related quality of life in the Cambridge City over-75s Cohort (CC75C): development of a dementia-specific scale and descriptive analyses.

BACKGROUND: The assessment of Health Related Quality of Life (HRQL) is important in people with dementia as it could influence their care and support plan. Many studies on dementia do not specifically set out to measure dementia-specific HRQL but do include related items. The aim of this study is to explore the distribution of HRQL by functional and socio-demographic variables in a population-based setting. METHODS: Domains of DEMQOL's conceptual framework were mapped in the Cambridge City over 75's Cohort (CC75C) Study. HRQL was estimated in 110 participants aged 80+ years with a confirmed diagnosis of dementia with mild/moderate severity. Acceptability (missing values and normality of the total score), internal consistency (Cronbach's alpha), convergent, discriminant and known group differences validity (Spearman correlations, Wilcoxon Mann-Whitney and Kruskal-Wallis tests) were assessed. The distribution of HRQL by socio-demographic and functional descriptors was explored. RESULTS: The HRQL score ranged from 0 to 16 and showed an internal consistency Alpha of 0.74. Validity of the instrument was found to be acceptable. Men had higher HRQL than women. Marital status had a greater effect on HRQL for men than it did for women. The HRQL of those with good self-reported health was higher than those with fair/poor self-reported health. HRQL was not associated with dementia severity. CONCLUSIONS: To our knowledge this is the first study to examine the distribution of dementia-specific HRQL in a population sample of the very old. We have mapped an existing conceptual framework of dementia specific HRQL onto an existing study and demonstrated the feasibility of this approach. Findings in this study suggest that whereas there is big emphasis in dementia severity, characteristics such as gender should be taken into account when assessing and implementing programmes to improve HRQL.