Endosymbiosis in trypanosomatids: the genomic cooperation between bacterium and host in the synthesis of essential amino acids is heavily influenced by multiple horizontal gene transfers

Background Trypanosomatids of the genera Angomonas and Strigomonas live in a mutualistic association characterized by extensive metabolic cooperation with obligate endosymbiotic Betaproteobacteria. However, the role played by the symbiont has been more guessed by indirect means than evidenced. Symbiont-harboring trypanosomatids, in contrast to their counterparts lacking symbionts, exhibit lower nutritional requirements and are autotrophic for essential amino acids. To evidence the symbiont’s contributions to this autotrophy, entire genomes of symbionts and trypanosomatids with and without symbionts were sequenced here. Results Analyses of the essential amino acid pathways revealed that most biosynthetic routes are in the symbiont genome. By contrast, the host trypanosomatid genome contains fewer genes, about half of which originated from different bacterial groups, perhaps only one of which (ornithine cyclodeaminase, EC:4.3.1.12) derived from the symbiont. Nutritional, enzymatic, and genomic data were jointly analyzed to construct an integrated view of essential amino acid metabolism in symbiont-harboring trypanosomatids. This comprehensive analysis showed perfect concordance among all these data, and revealed that the symbiont contains genes for enzymes that complete essential biosynthetic routes for the host amino acid production, thus explaining the low requirement for these elements in symbiont-harboring trypanosomatids. Phylogenetic analyses show that the cooperation between symbionts and their hosts is complemented by multiple horizontal gene transfers, from bacterial lineages to trypanosomatids, that occurred several times in the course of their evolution. Transfers occur preferentially in parts of the pathways that are missing from other eukaryotes. Conclusion We have herein uncovered the genetic and evolutionary bases of essential amino acid biosynthesis in several trypanosomatids with and without endosymbionts, explaining and complementing decades of experimental results. We uncovered the remarkable plasticity in essential amino acid biosynthesis pathway evolution in these protozoans, demonstrating heavy influence of horizontal gene transfer events, from Bacteria to trypanosomatid nuclei, in the evolution of these pathways.

A proteomic analysis of the functional effects of fatty acids in NIH 3T3 fibroblasts

Previous studies have demonstrated that long chain fatty acids influence fibroblast function at sub-lethal concentrations. This study is the first to assess the effects of oleic, linoleic or palmitic acids on protein expression of fibroblasts, as determined by standard proteomic techniques. The fatty acids were not cytotoxic at the concentration used in this work as assessed by membrane integrity, DNA fragmentation and the MTT assay but significantly increased cell proliferation. Subsequently, a proteomic analysis was performed using two dimensional difference gel electrophoresis (2D-DIGE) and MS based identification. Cells treated with 50 μM oleic, linoleic or palmitic acid for 24 h were associated with 24, 22, 16 spots differentially expressed, respectively. Among the identified proteins, α-enolase and far upstream element binding protein 1 (FBP-1) are of importance due to their function in fibroblast-associated diseases. However, modulation of α-enolase and FBP-1 expression by fatty acids was not validated by the Western blot technique.

Mechanisms underlying skeletal muscle insulin resistance induced by fatty acids: importance of the mitochondrial function

Insulin resistance condition is associated to the development of several syndromes, such as obesity, type 2 diabetes mellitus and metabolic syndrome. Although the factors linking insulin resistance to these syndromes are not precisely defined yet, evidence suggests that the elevated plasma free fatty acid (FFA) level plays an important role in the development of skeletal muscle insulin resistance. Accordantly, in vivo and in vitro exposure of skeletal muscle and myocytes to physiological concentrations of saturated fatty acids is associated with insulin resistance condition. Several mechanisms have been postulated to account for fatty acids-induced muscle insulin resistance, including Randle cycle, oxidative stress, inflammation and mitochondrial dysfunction. Here we reviewed experimental evidence supporting the involvement of each of these propositions in the development of skeletal muscle insulin resistance induced by saturated fatty acids and propose an integrative model placing mitochondrial dysfunction as an important and common factor to the other mechanisms.

Potential therapeutic effects of branched-chain amino acids supplementation on resistance exercise-based muscle damage in humans

Branched-chain amino acids (BCAA) supplementation has been considered an interesting nutritional strategy to improve skeletal muscle protein turnover in several conditions. In this context, there is evidence that resistance exercise (RE)-derived biochemical markers of muscle soreness (creatine kinase (CK), aldolase, myoglobin), soreness, and functional strength may be modulated by BCAA supplementation in order to favor of muscle adaptation. However, few studies have investigated such effects in well-controlled conditions in humans. Therefore, the aim of this short report is to describe the potential therapeutic effects of BCAA supplementation on RE-based muscle damage in humans. The main point is that BCAA supplementation may decrease some biochemical markers related with muscle soreness but this does not necessarily reflect on muscle functionality.

Absorption and metabolism of volatile fatty acids by rumen and omasum

Volatile fatty acids (VFA) absorption and metabolic capacity of rumen and omasum were compared, in vitro. Fragments of rumen wall and omasum laminae were taken from eight adult crossbred bovines. An isolated fragment of the mucosa was fitted in a tissue diffusion chamber. Valeric acid and CrEDTA were added to ruminal fluid and placed on the mucosal side and buffer solution was placed on the serosal side. Fractional absorption rates were measured by exponential VFA:Cr ratio decay over time. Metabolism rate was determined as the difference between VFA absorbed and VFA which appeared on the serosal side over time. Mitotic index was higher in omasum (0.52%) than in rumen epithelium (0.28%). VFA fractional absorption rate was higher in omasum (4.6%/h.cm²) than in rumen (0.4%/h.cm²). Acetate, propionate, butyrate, and valerate showed similar fractional absorption rates in both fragments. Percentage of metabolized acetate and propionate was lower than butyrate and valerate in both stomach compartments. In the rumen, individual VFA metabolism rates were similar (mean of 7.7 , but in the omasum, valerate (90.0 was more metabolized than butyrate (59.6 propionate (69.8 and acetate (51.7 . Correlation between VFA metabolism and mitotic index was positive in the rumen and in the omasum. In conclusion, VFA metabolism and absorption potential per surface of the omasum is higher than that of the rumen. Variations on rumen and omasum absorption capacities occur in the same way, and there are indications that factors capable of stimulating rumen wall proliferation are similarly capable of stimulating omasum walls.

Importance of the proportions of dietary polyunsaturated fatty acids and antioxoidants in larval development of marine fish

La acuicultura es uno de los sectores con mayor crecimiento en la producción animal mundial, con una tasa superior al 5% anual en la última década (Izquierdo et al., 2008). En la acuicultura, el éxito del cultivo de cualquier especie de peces marinos está limitado por la cantidad y la calidad de la producción en masa de sus alevines (Izquierdo y Fernández - Palacios, 1997). Por lo tanto, para cubrir las crecientes demandas de la producción de semillas, es necesario mejorar la calidad nutritiva de sus larvas, lo cual todavía constituye una de las principales limitaciones para el desarrollo del cultivo de especies de peces marinos (Watanabe et al., 1983; Yúfera y Pascual, 1984; Sargent et al. 1997; Izquierdo et al., 2000). Aunque la producción Mediterránea del cultivo de peces marinos se ha incrementado en varias especies, la dorada Sparus aurata continúa siendo la especie mas cultivada (Izquierdo, 2005), con una producción anual de alevines que supera los 120 millones/ año. La demanda de alevines de buena calidad está aumentando a un ritmo del 10% anual, pero el éxito de la producción de los juveniles se ve muy afectado por la eficacia de la primera alimentación y la calidad nutricional de la dieta de arranque (Kolkovski et al., 1993; Sargent et al., 1997; Izquierdo et al., 2000). En general, la dorada y la lubina europea (Dicentrarchus labrax) son las especies más importantes de peces marinos criados en la región mediterránea y han caracterizado el desarrollo de la acuicultura marina en esta región en las últimas tres décadas (FAO, 1999). Además, en la producción, de ambas especies, se prevee una mayor expansión (Basurco y Abellán, 1999). Sin embargo, a pesar de que el engorde de estas especies está bien controlado, el conocimiento de sus necesidades nutricionales, en comparación con otras especies como salmónidos y carpas, sigue siendo incompleto (NRC, 1993). Por lo tanto, para obtener un mejor crecimiento y una mayor tasa de supervivencia, es esencial una dieta que responda a las necesidades nutricionales de las larvas, tanto cualitativas como cuantitativas (Kolkovski et al., 1993; Sargent et al., 1997). Además, incluso antes de que comience el desarrollo larvario, los huevos de los peces deben contener todos los nutrientes que cubran los requerimientos adecuados para el desarrollo del embrión (Izquierdo y Fernández - Palacios, 1997). En los últimos años, las investigaciones han prestado gran atención a la importancia de los lípidos dietéticos para larvas de peces marinos (Izquierdo et al., 2003), puesto que son esenciales para el crecimiento y el desarrollo de los mismos (Watanabe, 1982; Sargent et al., 1999a). Como reflejo de esta importancia, la dieta para dorada y lubina se ha convertido en una dieta altamente energética (25% de lípidos) en comparación con la década anterior (12% de lípidos) (Izquierdo et al., 2003). Los lípidos dietéticos proporcionan una fuente rica de energía y fosfolípidos y son fundamentales para la estructura de las biomembranas. Los lípidos dietéticos también sirven como vehículos para la absorción de otros nutrientes, como las vitaminas liposolubles A, D, E, y K, y pigmentos naturales o sintéticos. Los lípidos son componentes de las hormonas y precursores para la síntesis de diversos metabolitos funcionales, como las prostaglandinas y otros eicosonoides. Además, los lípidos dietéticos son reconocidos como uno de los factores nutricionales más importantes que afectan el crecimiento y la supervivencia de las larvas (Watanabe et al., 1983), porque constituyen materiales esenciales para la formación normal de célula, las membranas de los tejidos y el desarrollo de órganos (Izquierdo et al., 1998, 2003; Pousaò et al., 2003). Sin embargo, la utilización de los lípidos dietéticos por las larvas puede verse afectada directa o indirectamente por varios cambios morfológicos y fisiológicos que ocurren durante el desarrollo larvario. En los últimos años ha habido más interés en todos estos aspectos nutricionales de los lípidos en las larvas de peces, debido a la importancia de utilización de los lípidos dieteticos para el óptimo crecimiento y supervivencia larvaria (Izquierdo et al., 2000).

Effects of dietary fatty acids on patterns of early mineralisation and bone formation in the axial skeleton of Sparus aurata (Linnaeus, 1758)

Máster Oficial en Cultivos Marinos. VI Máster Internacional en Acuicultura. Trabajo presentado como requisito parcial para la obtención del Título de Máster Oficial en Cultivos Marinos, otorgado por la Universidad de Las Palmas de Gran Canaria (ULPGC), el Instituto Canario de Ciencias Marinas (ICCM), y el Centro Internacional de Altos Estudios Agronómicos Mediterráneos de Zaragoza (CIHEAM)

Enhancement of gilthead sea bream (Sparus aurata) and sea bass (Dicentrarchus labrax) larval growth by dietary vitamin E en relation to diffetent levels of essential fatty acids

[EN]Most marine fish larvae require high amounts of n-3 HUFA (highly unsaturated fatty acids) such as eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) (Watanabe, 1982; Izquierdo, 1996). Fish larvae tissue lipids are also very high in n-3 HUFA, what implies a higher risk of peroxidation (Sargent et al. 1999) and cellular damage (Kanazawa, 1991), requiring then antioxidants to protect them intra- and extra-cellularly from free radical compounds. Vitamin E (Vit E) functions as a chain breaking antioxidant, reacting with the lipid peroxide radical produced and preventing the further reaction with a new PUFA. Hence their requirements are related with the dietary and tissue PUFA contents. The objective of the present study was to determine the effect of dietary Vit E on gilthead sea bream and sea bass survival, growth and stress, at different n-3 HUFA levels.

Synthetic and mechanistic investigation of new radical processes: reaction of organic azides with group-XIII Lewis acids

Dichloroindium hydride revealed to be a valid alternative to tributyltin hydride for radical reduction of organic (alkyl, aryl, acyl, solfonyl) azides. The new approach entails mild reaction conditions and provides high yields of the corresponding amines and amides, also showing high degrees of selectivity. The system dichloroindium hydride / azides can be utilised in fivemembered ring closures of g-azidonitriles, as a new source of aminyl radicals for the attractive synthesis of interesting amidine compounds in the absence of both toxic reagents and tedious purification procedures. Allylindium dichloride seems a good substitute for dichloroindium hydride for generation of indium centred radicals under photolytic conditions, since it allows allylation of electrophilic azides (e.g. phenylsulfonyl azide) and halogen or ester δ-substituted azides, the latter through a 1,5-H transfer rearrangement mechanism. Evidences of the radical nature of the reactions mechanism were provided by ESR spectroscopy, furthermore the same technique, allowed to discover that the reaction of azides with indium trichloride and other group XIII Lewis acids, in particular gallium trichloride, gives rise to strongly coloured, persistent paramagnetic species, whose structure is consistent with the radical cation of the head-to-tail dimer of the aniline corresponding to the starting azide.

First evidences of the effect of polyunsaturated fatty acids in fish neural activity and their implications in behaviour

Programa de doctorado: Acuicultura: producción controlada de animales acuáticos; Grupo de Investigación en Acuicultura (GIA)

N-3 fatty acids and cardiovascular prevention

In this study we elucidate the role of polyunsaturated fatty acids (PUFAs) in the prevention of cardiovascular diseases, focusing the attention on their role in the modulation of acyl composition of cell lipids and of gene expression. Regarding this latter mechanism, the effectiveness of PUFAs as activators of two transcriptional factors, SREBPs and PPARs, have been considered. Two different model system have been used: primary cultures of neonatal rat cardiomyocytes and an human hepatoma cell line (HepG2). Cells have been supplemented with different PUFAs at physiological concentration, and special attention has been devoted to the main n-3 PUFAs, EPA and DHA. PUFAs influence on global gene expression in cardiomyocytes has been evaluated using microarray technique. Furthermore, since it is not fully elucidated which transcription factors are involved in this modulation in the heart, expression and activation of the three different PPAR isoforms have been investigated. Hepatocytes have been used as experimental model system in the evaluation of PUFAs effect on SREBP activity. SREBPs are considered the main regulator of cholesterol and triglyceride synthesis, which occur mainly in the liver. In both experimental models the modification of cell lipid fatty acid composition subsequent to PUFAs supplementation has been evaluated, and related to the effects observed at molecular level. The global vision given by the obtained results may be important for addressing new researches and be useful to educators and policy makers in setting recommendations for reaching optimal health through good nutrition.