Search

Abstract Background The European sea bass (Dicentrarchus labrax) is a marine fish of great importance for fisheries and aquaculture. Functional genomics offers the possibility to discover the molecular mechanisms underlying productive traits in farmed fish, and a step towards the application of marker assisted selection methods in this species. To this end, we report here on the development of an oligo DNA microarray for D. labrax. Results A database consisting of 19,048 unique transcripts was constructed, of which 12,008 (63%) could be annotated by similarity and 4,692 received a GO functional annotation. Two non-overlapping 60mer probes were designed for each unique transcript and in-situ synthesized on glass slides using Agilent SurePrint™ technology. Probe design was positively completed for 19,035 target clusters; the oligo microarray was then applied to profile gene expression in mandibles and whole-heads of fish affected by prognathism, a skeletal malformation that strongly affects sea bass production. Statistical analysis identified 242 transcripts that are significantly down-regulated in deformed individuals compared to normal fish, with a significant enrichment in genes related to nervous system development and functioning. A set of genes spanning a wide dynamic range in gene expression level were selected for quantitative RT-PCR validation. Fold change correlation between microarray and qPCR data was always significant. Conclusions The microarray platform developed for the European sea bass has a high level of flexibility, reliability, and reproducibility. Despite the well known limitations in achieving a proper functional annotation in non-model species, sufficient information was obtained to identify biological processes that are significantly enriched among differentially expressed genes. New insights were obtained on putative mechanisms involved on mandibular prognathism, suggesting that bone/nervous system development might play a role in this phenomenon.

Abstract Background Aquaculture represents the most sustainable alternative of seafood supply to substitute for the declining marine fisheries, but severe production bottlenecks remain to be solved. The application of genomic technologies offers much promise to rapidly increase our knowledge on biological processes in farmed species and overcome such bottlenecks. Here we present an integrated platform for mRNA expression profiling in the gilthead sea bream (Sparus aurata), a marine teleost of great importance for aquaculture. Results A public data base was constructed, consisting of 19,734 unique clusters (3,563 contigs and 16,171 singletons). Functional annotation was obtained for 8,021 clusters. Over 4,000 sequences were also associated with a GO entry. Two 60mer probes were designed for each gene and in-situ synthesized on glass slides using Agilent SurePrint™ technology. Platform reproducibility and accuracy were assessed on two early stages of sea bream development (one-day and four days old larvae). Correlation between technical replicates was always > 0.99, with strong positive correlation between paired probes. A two class SAM test identified 1,050 differentially expressed genes between the two developmental stages. Functional analysis suggested that down-regulated transcripts (407) in older larvae are mostly essential/housekeeping genes, whereas tissue-specific genes are up-regulated in parallel with the formation of key organs (eye, digestive system). Cross-validation of microarray data was carried out using quantitative qRT-PCR on 11 target genes, selected to reflect the whole range of fold-change and both up-regulated and down-regulated genes. A statistically significant positive correlation was obtained comparing expression levels for each target gene across all biological replicates. Good concordance between qRT-PCR and microarray data was observed between 2- and 7-fold change, while fold-change compression in the microarray was present for differences greater than 10-fold in the qRT-PCR. Conclusion A highly reliable oligo-microarray platform was developed and validated for the gilthead sea bream despite the presently limited knowledge of the species transcriptome. Because of the flexible design this array will be able to accommodate additional probes as soon as novel unique transcripts are available.

Abstract Background Oceans cover approximately 70% of the Earth's surface with an average depth of 3800 m and a pressure of 38 MPa, thus a large part of the biosphere is occupied by high pressure environments. Piezophilic (pressure-loving) organisms are adapted to deep-sea life and grow optimally at pressures higher than 0.1 MPa. To better understand high pressure adaptation from a genomic point of view three different Photobacterium profundum strains were compared. Using the sequenced piezophile P. profundum strain SS9 as a reference, microarray technology was used to identify the genomic regions missing in two other strains: a pressure adapted strain (named DSJ4) and a pressure-sensitive strain (named 3TCK). Finally, the transcriptome of SS9 grown under different pressure (28 MPa; 45 MPa) and temperature (4°C; 16°C) conditions was analyzed taking into consideration the differentially expressed genes belonging to the flexible gene pool. Results These studies indicated the presence of a large flexible gene pool in SS9 characterized by various horizontally acquired elements. This was verified by extensive analysis of GC content, codon usage and genomic signature of the SS9 genome. 171 open reading frames (ORFs) were found to be specifically absent or highly divergent in the piezosensitive strain, but present in the two piezophilic strains. Among these genes, six were found to also be up-regulated by high pressure. Conclusion These data provide information on horizontal gene flow in the deep sea, provide additional details of P. profundum genome expression patterns and suggest genes which could perform critical functions for abyssal survival, including perhaps high pressure growth.

Background: Whole genome and exome sequencing are contributing to the extraordinary progress in the study of human genetic variants. In this fast developing field, appropriate and easily accessible tools are required to facilitate data analysis. Results: Here we describe QueryOR, a web platform suitable for searching among known candidate genes as well as for finding novel gene-disease associations. QueryOR combines several innovative features that make it comprehensive, flexible and easy to use. Instead of being designed on specific datasets, it works on a general XML schema specifying formats and criteria of each data source. Thanks to this flexibility, new criteria can be easily added for future expansion. Currently, up to 70 user-selectable criteria are available, including a wide range of gene and variant features. Moreover, rather than progressively discarding variants taking one criterion at a time, the prioritization is achieved by a global positive selection process that considers all transcript isoforms, thus producing reliable results. QueryOR is easy to use and its intuitive interface allows to handle different kinds of inheritance as well as features related to sharing variants in different patients. QueryOR is suitable for investigating single patients, families or cohorts. Conclusions: QueryOR is a comprehensive and flexible web platform eligible for an easy user-driven variant prioritization. It is freely available for academic institutions at http://queryor.cribi.unipd.it/. [ABSTRACT FROM AUTHOR]

Background: We performed an analysis of the transcriptome during the blastogenesis of the chordate Botryllus schlosseri, focusing in particular on genes involved in cell death by apoptosis. The tunicate B. schlosseri is an ascidian forming colonies characterized by the coexistence of three blastogenetic generations: filter-feeding adults, buds on adults, and budlets on buds. Cyclically, adult tissues undergo apoptosis and are progressively resorbed and replaced by their buds originated by asexual reproduction. This is a feature of colonial tunicates, the only known chordates that can reproduce asexually. Results: Thanks to a newly developed web-based platform (http://botryllus.cribi.unipd.it), we compared the transcriptomes of the mid-cycle, the pre-take-over, and the take-over phases of the colonial blastogenetic cycle. The platform is equipped with programs for comparative analysis and allows to select the statistical stringency. We enriched the genome annotation with 11,337 new genes; 581 transcripts were resolved as complete open reading frames, translated in silico into amino acid sequences and then aligned onto the non-redundant sequence database. Significant differentially expressed genes were classified within the gene ontology categories. Among them, we recognized genes involved in apoptosis activation, de-activation, and regulation. Conclusions: With the current work, we contributed to the improvement of the first released B. schlosseri genome assembly and offer an overview of the transcriptome changes during the blastogenetic cycle, showing up- and down-regulated genes. These results are important for the comprehension of the events underlying colony growth and regression, cell proliferation, colony homeostasis, and competition among different generations. [ABSTRACT FROM AUTHOR]

Photobacterium profundum is a cosmopolitan marine bacterium capable of growth at low temperature and high hydrostatic pressure. Multiple strains of P. profundum have been isolated from different depths of the ocean and display remarkable differences in their physiological responses to pressure. The genome sequence of the deep-sea piezopsychrophilic strain Photobacterium profundum SS9 has provided some clues regarding the genetic features required for growth in the deep sea. The sequenced genome of Photobacterium profundum strain 3TCK, a non-piezophilic strain isolated from a shallow-water environment, is now available and its analysis expands the identification of unique genomic features that correlate to environmental differences and define the Hutchinsonian niche of each strain. These differences range from variations in gene content to specific gene sequences under positive selection. Genome plasticity between Photobacterium bathytypes was investigated when strain 3TCK-specific genes involved in photorepair were introduced to SS9, demonstrating that horizontal gene transfer can provide a mechanism for rapid colonisation of new environments. [ABSTRACT FROM AUTHOR]

ZASP is a cytoskeletal PDZ-LIM protein predominantly expressed in striated muscle. It forms multiprotein complexes and plays a pivotal role in the structural integrity of sarcomeres. Mutations in the ZASP protein are associated with myofibrillar myopathy, left ventricular non-compaction and dilated cardiomyopathy. The ablation of its murine homologue Cypher results in neonatal lethality. ZASP has several alternatively spliced isoforms, in this paper we clarify the nomenclature of its human isoforms as well as their dynamics and expression pattern in striated muscle. Interaction is demonstrated between ZASP and two new binding partners both of which have roles in signalling, regulation of gene expression and muscle differentiation; the mechanosensing protein Ankrd2 and the tumour suppressor protein p53. These proteins and ZASP form a triple complex that appears to facilitate poly-SUMOylation of p53. We also show the importance of two of its functional domains, the ZM-motif and the PDZ domain. The PDZ domain can bind directly to both Ankrd2 and p53 indicating that there is no competition between it and p53 for the same binding site on Ankrd2. However there is competition for this binding site between p53 and a region of the ZASP protein lacking the PDZ domain, but containing the ZM-motif. ZASP is negative regulator of p53 in transactivation experiments with the p53-responsive promoters, MDM2 and BAX. Mutations in the ZASP ZM-motif induce modification in protein turnover. In fact, two mutants, A165V and A171T, were not able to bind Ankrd2 and bound only poorly to alpha-actinin2. This is important since the A165V mutation is responsible for zaspopathy, a well characterized autosomal dominant distal myopathy. Although the mechanism by which this mutant causes disease is still unknown, this is the first indication of how a ZASP disease associated mutant protein differs from that of the wild type ZASP protein. [ABSTRACT FROM AUTHOR]

The horse is an optimal model organism for studying the genomic response to exercise-induced stress, due to its natural aptitude for athletic performance and the relative homogeneity of its genetic and environmental backgrounds. Here, we applied RNA-sequencing analysis through the use of SOLiD technology in an experimental framework centered on exercise-induced stress during endurance races in equine athletes. We monitored the transcriptional landscape by comparing gene expression levels between animals at rest and after competition. Overall, we observed a shift from coding to non-coding regions, suggesting that the stress response involves the differential expression of not annotated regions. Notably, we observed significant post-race increases of reads that correspond to repeats, especially the intergenic and intronic L1 and L2 transposable elements. We also observed increased expression of the antisense strands compared to the sense strands in intronic and regulatory regions (1 kb up- and downstream) of the genes, suggesting that antisense transcription could be one of the main mechanisms for transposon regulation in the horse under stress conditions. We identified a large number of transcripts corresponding to intergenic and intronic regions putatively associated with new transcriptional elements. Gene expression and pathway analysis allowed us to identify several biological processes and molecular functions that may be involved with exercise-induced stress. Ontology clustering reflected mechanisms that are already known to be stress activated (e.g., chemokine-type cytokines, Toll-like receptors, and kinases), as well as “nucleic acid binding” and “signal transduction activity” functions. There was also a general and transient decrease in the global rates of protein synthesis, which would be expected after strenuous global stress. In sum, our network analysis points toward the involvement of specific gene clusters in equine exercise-induced stress, including those involved in inflammation, cell signaling, and immune interactions. [ABSTRACT FROM AUTHOR]

Wheat is one of the world's most important crops and is characterized by a large polyploid genome. One way to reduce genome complexity is to isolate single chromosomes using flow cytometry. Low coverage DNA sequencing can provide a snapshot of individual chromosomes, allowing a fast characterization of their main features and comparison with other genomes. We used massively parallel 454 pyrosequencing to obtain a 2x coverage of wheat chromosome 5A. The resulting sequence assembly was used to identify TEs, genes and miRNAs, as well as to infer a virtual gene order based on the synteny with other grass genomes. Repetitive elements account for more than 75% of the genome. Gene content was estimated considering non-redundant reads showing at least one match to ESTs or proteins. The results indicate that the coding fraction represents 1.08% and 1.3% of the short and long arm respectively, projecting the number of genes of the whole chromosome to approximately 5,000. 195 candidate miRNA precursors belonging to 16 miRNA families were identified. The 5A genes were used to search for syntenic relationships between grass genomes. The short arm is closely related to Brachypodium chromosome 4, sorghum chromosome 8 and rice chromosome 12; the long arm to regions of Brachypodium chromosomes 4 and 1, sorghum chromosomes 1 and 2 and rice chromosomes 9 and 3. From these similarities it was possible to infer the virtual gene order of 392 (5AS) and 1,480 (5AL) genes of chromosome 5A, which was compared to, and found to be largely congruent with the available physical map of this chromosome. [ABSTRACT FROM AUTHOR]

The analysis of the first plant genomes provided unexpected evidence for genome duplication events in species that had previously been considered as true diploids on the basis of their genetics. These polyploidization events may have had important consequences in plant evolution, in particular for species radiation and adaptation and for the modulation of functional capacities. Here we report a high-quality draft of the genome sequence of grapevine (Vitis vinifera) obtained from a highly homozygous genotype. The draft sequence of the grapevine genome is the fourth one produced so far for flowering plants, the second for a woody species and the first for a fruit crop (cultivated for both fruit and beverage). Grapevine was selected because of its important place in the cultural heritage of humanity beginning during the Neolithic period. Several large expansions of gene families with roles in aromatic features are observed. The grapevine genome has not undergone recent genome duplication, thus enabling the discovery of ancestral traits and features of the genetic organization of flowering plants. This analysis reveals the contribution of three ancestral genomes to the grapevine haploid content. This ancestral arrangement is common to many dicotyledonous plants but is absent from the genome of rice, which is a monocotyledon. Furthermore, we explain the chronology of previously described whole-genome duplication events in the evolution of flowering plants. [ABSTRACT FROM AUTHOR]

Abstract: Two-thirds of Earth''s surface is covered by oceans, yet the study of this massive integrated living system is still in its infancy. Various environmental variables, such as high salinity, low and changeable nutrient availability and depth-correlated gradients of light, temperature, nutrients and pressure shape the diversity, physiology and ecology of marine species. As oceans present an average depth of 3800m, deep-sea ecosystems represent the most common marine ecological niche. One of the key environment variables that influences the life and evolution of deep-sea organisms is high pressure. This extreme widespread condition requires specific adaptations, the nature of which remains largely unknown. Recent advances in genomic approaches, such as in sequencing technologies and global expression profiling, are rapidly increasing the data available to understand microbial evolution, biochemistry, physiology and diversity. This review summarises the analysis of the results published so far about microbial high pressure adaptation from a genomic point of view. Understanding high pressure adaptation mechanisms is not just a scientific exercise but has important biotechnological implications. For example, hydrostatic pressure is a reality for food science and technology, both for food preparation and preservation. An understanding of the effects of pressure on biomolecules will expand its use in the medical, industrial and biotechnological fields. [Copyright &y& Elsevier]