An EST resource for tilapia based on 17 normalized libraries and assembly of 116,899 sequence tags

Background

Large collections of expressed sequence tags (ESTs) are a fundamental resource for analysis of gene expression and annotation of genome sequences. We generated 116,899 ESTs from 17 normalized and two non-normalized cDNA libraries representing 16 tissues from tilapia, a cichlid fish widely used in aquaculture and biological research.

Results

The ESTs were assembled into 20,190 contigs and 36,028 singletons for a total of 56,218 unique sequences and a total assembled length of 35,168,415 bp. Over the whole project, a unique sequence was discovered for every 2.079 sequence reads. 17,722 (31.5%) of these unique sequences had significant BLAST hits (e-value < 10^-10) to the UniProt database.

Conclusion

Normalization of the cDNA pools with double-stranded nuclease allowed us to efficiently sequence a large collection of ESTs. These sequences are an important resource for studies of gene expression, comparative mapping and annotation of the forthcoming tilapia genome sequence.

     

The MAR1 transporter is an opportunistic entry point for antibiotics

The vast quantities of antibiotics used in modern agriculture contaminate the environment and threaten human health. Recent studies have shown that crop plants grown in soil fertilized with manure from antibiotic-treated animals can accumulate antibiotic within the plant body, thus making them an additional antibiotic exposure route for consumers. Until recently, mechanisms of antibiotic entry and subcellular partitioning within plant cells were virtually unknown. We have uncovered and characterized a transporter gene in Arabidopsis thaliana, MAR1, which appears to control antibiotic entry into the chloroplast. Antibiotic resistance via MAR1 is specific to the aminoglycoside class, and is conferred by loss-of-function mutations, which is rather unusual, since most transporter-based antibiotic resistance is conferred by overexpression or gain-of-function mutations in efflux pumps with poor substrate specificity. Since MAR1 overexpression lines exhibit various iron starvation phenotypes, we propose that MAR1 transports an iron chelation molecule that is mimicked specifically by aminoglycoside antibiotics, and this facilitates their entry into the chloroplast. Knowledge about MAR1 enhances our understanding of how antibiotics might enter the plant cell, which may aid in the production of crop plants that are incapable of antibiotic accumulation, as well as further the development of new plant-based antibiotic resistance markers.Key words: antibiotic, contamination, transport, import, chloroplast, membrane, iron, chelation, nicotianamine     

Diet effects on honeybee immunocompetence

The maintenance of the immune system can be costly, and a lack of dietary protein can increase the susceptibility of organisms to disease. However, few studies have investigated the relationship between protein nutrition and immunity in insects. Here, we tested in honeybees (Apis mellifera) whether dietary protein quantity (monofloral pollen) and diet diversity (polyfloral pollen) can shape baseline immunocompetence (IC) by measuring parameters of individual immunity (haemocyte concentration, fat body content and phenoloxidase activity) and glucose oxidase (GOX) activity, which enables bees to sterilize colony and brood food, as a parameter of social immunity. Protein feeding modified both individual and social IC but increases in dietary protein quantity did not enhance IC. However, diet diversity increased IC levels. In particular, polyfloral diets induced higher GOX activity compared with monofloral diets, including protein-richer diets. These results suggest a link between protein nutrition and immunity in honeybees and underscore the critical role of resource availability on pollinator health.     

The MitoDrome database annotates and compares the OXPHOS nuclear genes of Drosophila melanogaster, Drosophila pseudoobscura and Anopheles gambiae

The oxidative phosphorylation (OXPHOS) is the primary energy-producing process of all aerobic organisms and the only cellular function under the dual control of both the mitochondrial and the nuclear genomes. Functional characterization and evolutionary study of the OXPHOS system is of great importance for the understanding of many as yet unclear aspects of nucleus-mitochondrion genomic co-evolution and co-regulation gene networks. The MitoDrome database is a web-based database which provides genomic annotations about nuclear genes of Drosophila melanogaster encoding for mitochondrial proteins. Recently, MitoDrome has included a new section annotating genomic information about OXPHOS genes in Drosophila pseudoobscura and Anopheles gambiae and their comparative analysis with their Drosophila melanogaster and human counterparts. The introduction of this new comparative annotation section into MitoDrome is expected to be a useful resource for both functional and structural genomics related to the OXPHOS system.     

Worker-worker inhibition of honey bee behavioural development independent of queen and brood

Summary. Foragers inhibit the behavioural development of young adult worker honey bees, delaying the age at onset of foraging. But the similar effect caused by pheromones produced by both the queen and brood raised the possibility that some of the previously attributed forager effects might be due to queen, brood, or both. Here we studied whether physical contacts between young bees and old foragers can inhibit behavioural development while controlling for queen and brood effects. Results demonstrated that foragers inhibit the behavioural development of young adult worker bees independent of the queen and brood, via a mechanism that requires physical contact.Received 24 November 2003; revised 27 March 2004; accepted 21 April 2004.     

Genetic diversity and recruitment of the tropical palm, Euterpe edulis Mart., in a natural population from the Brazilian Atlantic Forest

The genetic diversity and recruitment of plants of heart-of-palm tree (Euterpe edulis Mart.) were investigated in a natural population located in Southern Brazil. Five categories of plants, from seedlings to adults, were analysed using 16 allozymic loci. The results showed an average population level of genetic diversity (He=0.278) greater than the average of plant species already studied. The recruitment process of E. edulis is related to its genetic characteristics. A significant increase in the heterozygote frequency towards the adult stages was observed at three loci (Pgdh-2, G6pdh-1 and Mdh-1). This suggests the possible action of natural selection in promoting such heterozygote increase. In the same way, a linear increase in allele frequencies was observed at four loci (Prx-3, Prx-4, Pgdh-2 and G6pdh-1), indicating that recruitment is also related to a greater survival of individuals that are carriers of certain alleles. The maintenance of high diversity levels, as well as the increase in the heterozygote frequency, are positive aspects for in situ conservation. Furthermore, an increase in heterozygote frequency is favourable to the management of the species, since the maintenance of a stock of reproductive individuals with high heterozygosity levels favours the maintenance of the population dynamic and structure.     

Differential effects of coconut oil- and fish oil-enriched diets on tricarboxylate carrier in rat liver mitochondria

The mitochondrial tricarboxylate carrier (TCC) plays an important role in lipogenesis being TCC-responsible for the efflux from the mitochondria to the cytosol of acetyl-CoA, the primer for fatty acid synthesis. In this study, we investigated the effects of two high-fat diets with different fatty acid composition on the hepatic TCC activity. Rats were fed for 3 weeks on a basal diet supplemented with 15% of either coconut oil (CO), abundant in medium-chain saturated fatty acids, or fish oil (FO), rich in n-3 polyunsaturated fatty acids. Mitochondrial fatty acid composition was differently influenced by the dietary treatments, while no appreciable change in phospholipid composition and cholesterol level was observed. Compared with CO, the TCC activity was markedly decreased in liver mitochondria from FO-fed rats; kinetic analysis of the carrier revealed a decrease of the Vmax, with no change of the Km. No difference in the Arrhenius plot between the two groups was observed. Interestingly, the carrier protein level and the corresponding mRNA abundance decreased following FO treatment. These data indicate that FO administration markedly decreased the TCC activity as compared with CO. This effect is most likely due to a reduced gene expression of the carrier protein.     

Characterization of DIP1, a novel nuclear protein in Drosophila melanogaster

We have recently identified in Drosophila melanogaster a new gene encoding a nuclear protein, DIP1. Here we report the developmental expression and the finding that DIP1 subcellular localization is in the nucleus and at the nuclear periphery during interphase in embryos. Interestingly, in humans, DIP1 antibody identified signals in nuclei from cultured cells and reacted with a rough 30kDa protein in Western blotting experiments, demonstrating evolutionary conservation.     

Solution structure of Apo Cu,Zn superoxide dismutase: role of metal ions in protein folding

The solution structure of the demetalated copper, zinc superoxide dismutase is obtained for the monomeric Glu133Gln/Phe50Glu/Gly51Glu mutant through NMR spectroscopy. The demetalated protein still has a well-defined tertiary structure; however, two beta-strands containing two copper ligands (His46 and His48, beta4) and one zinc ligand (Asp83, beta5) are shortened, and the sheet formed by these strands and strands beta7 and beta8 moves away from the other strands of the beta-barrel to form an open clam with respect to a closed conformation in the holoprotein. Furthermore, loop IV which contains three zinc ligands (His63, His71, and His80) and loop VII which contributes to the definition of the active cavity channel are severely disordered, and experience extensive mobility as it results from thorough (15)N relaxation measurements. These structural and mobility data, if compared with those of the copper-depleted protein and holoprotein, point out the role of each metal ion in the protein folding, leading to the final tertiary structure of the holoprotein, and provide hints for the mechanisms of metal delivery by metal chaperones.