An integrated genomic approach for the study of mandibular prognathism in the European seabass (Dicentrarchus labrax)

Skeletal anomalies in farmed fish are a relevant issue affecting animal welfare and health and causing significant economic losses. Here, a high-density genetic map of European seabass for QTL mapping of jaw deformity was constructed and a genome-wide association study (GWAS) was carried out on a total of 298 juveniles, 148 of which belonged to four full-sib families. Out of 298 fish, 107 were affected by mandibular prognathism (MP). Three significant QTLs and two candidate SNPs associated with MP were identified. The two GWAS candidate markers were located on ChrX and Chr17, both in close proximity with the peaks of the two most significant QTLs. Notably, the SNP marker on Chr17 was positioned within the Sobp gene coding region, which plays a pivotal role in craniofacial development. The analysis of differentially expressed genes in jaw-deformed animals highlighted the "nervous system development" as a crucial pathway in MP. In particular, Zic2, a key gene for craniofacial morphogenesis in model species, was significantly down-regulated in MP-affected animals. Gene expression data revealed also a significant down-regulation of Sobp in deformed larvae. Our analyses, integrating transcriptomic and GWA methods, provide evidence for putative mechanisms underlying seabass jaw deformity

Exploring the larval transcriptome of the common sole (Solea solea L.)

open7noBackground The common sole (Solea solea) is a promising candidate for European aquaculture; however, the limited knowledge of the physiological mechanisms underlying larval development in this species has hampered the establishment of successful flatfish aquaculture. Although the fact that genomic tools and resources are available for some flatfish species, common sole genomics remains a mostly unexplored field. Here, we report, for the first time, the sequencing and characterisation of the transcriptome of S. solea and its application for the study of molecular mechanisms underlying physiological and morphological changes during larval-to-juvenile transition. Results The S. solea transcriptome was generated from whole larvae and adult tissues using the Roche 454 platform. The assembly process produced a set of 22,223 Isotigs with an average size of 726 nt, 29 contigs and a total of 203,692 singletons. Of the assembled sequences, 75.2% were annotated with at least one known transcript/protein; these transcripts were then used to develop a custom oligo-DNA microarray. A total of 14,674 oligonucleotide probes (60 nt), representing 12,836 transcripts, were in situ synthesised onto the array using Agilent non-contact ink-jet technology. The microarray platform was used to investigate the gene expression profiles of sole larvae from hatching to the juvenile form. Genes involved in the ontogenesis of the visual system are up-regulated during the early stages of larval development, while muscle development and anaerobic energy pathways increase in expression over time. The gene expression profiles of key transcripts of the thyroid hormones (TH) cascade and the temporal regulation of the GH/IGF1 (growth hormone/insulin-like growth factor I) system suggest a pivotal role of these pathways in fish growth and initiation of metamorphosis. Pre-metamorphic larvae display a distinctive transcriptomic landscape compared to previous and later stages. Our findings highlighted the up-regulation of gene pathways involved in the development of the gastrointestinal system as well as biological processes related to folic acid and retinol metabolism. Additional evidence led to the formation of the hypothesis that molecular mechanisms of cell motility and ECM adhesion may play a role in tissue rearrangement during common sole metamorphosis. Conclusions Next-generation sequencing provided a good representation of the sole transcriptome, and the combination of different approaches led to the annotation of a high number of transcripts. The construction of a microarray platform for the characterisation of the larval sole transcriptome permitted the definition of the main processes involved in organogenesis and larval growth. Keywords: Solea solea; Flatfish; Larval development; Metamorphosis; Transcriptome; Gene expressionopenSerena Ferraresso; Alessio Bonaldo; Luca Parma; Stefano Cinotti; Paola Massi; Luca Bargelloni; Pier Paolo GattaSerena Ferraresso; Alessio Bonaldo; Luca Parma; Stefano Cinotti; Paola Massi; Luca Bargelloni; Pier Paolo Gatt

Array Comparative Genomic Hybridization Analysis Reveals Significantly Enriched Pathways in Canine Oral Melanoma

Human Mucosal Melanoma (hMM) is an aggressive neoplasm of neuroectodermal origin with distinctive features from the more common cutaneous form of malignant melanoma (cMM). At the molecular level, hMMs are characterized by large chromosomal aberrations rather than single-nucleotide mutations, typically observed in cMM. Given the scarcity of available cases, there have been many attempts to establish a reliable animal model. In pet dogs, Canine Oral Melanoma (COM) is the most common malignant tumor of the oral cavity, sharing clinical and histological aspects with hMM. To improve the knowledge about COM\u2019s genomic DNA alterations, in the present work, formalin-fixed, paraffin-embedded (FFPE) samples of COM from different European archives were collected to set up an array Comparative Genomic Hybridization (aCGH) analysis to estimate recurrent Copy Number Aberrations (CNAs). DNA was extracted in parallel from tumor and healthy fractions and 19 specimens were successfully submitted to labeling and competitive hybridization. Data were statistically analyzed through GISTIC2.0 and a pathway-enrichment analysis was performed with ClueGO. Recurrent gained regions were detected, affecting chromosomes CFA 10, 13 and 30, while lost regions involved chromosomes CFA 10, 11, 22, and 30. In particular, CFA 13 showed a whole-chromosome gain in 37% of the samples, while CFA 22 showed a whole-chromosome loss in 25%. A distinctive sigmoidal trend was observed in CFA 10 and 30 in 25 and 30% of the samples, respectively. Comparative analysis revealed that COM and hMM share common chromosomal changes in 32 regions. MAPK- and PI3K-related genes were the most frequently involved, while pathway analysis revealed statistically significant perturbation of cancer-related biological processes such as immune response, drug metabolism, melanocytes homeostasis, and neo-angiogenesis. The latter is a new evidence of a significant involvement of neovascularization-related pathways in COMs and can provide the rationale for future application in anti-cancer targeted therapies

Development of an oligo DNA microarray for the European sea bass and its application to expression profiling of jaw deformity

<p>Abstract</p> <p>Background</p> <p>The European sea bass (<it>Dicentrarchus labrax</it>) 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 <it>D. labrax</it>.</p> <p>Results</p> <p>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 <it>in-situ </it>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.</p> <p>Conclusions</p> <p>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.</p

Long Non-Coding RNAs as Molecular Signatures for Canine B-Cell Lymphoma Characterization

Background: Diffuse large B-cell lymphoma (DLBCL), marginal zone lymphoma (MZL) and follicular lymphoma (FL) are the most common B-cell lymphomas (BCL) in dogs. Recent investigations have demonstrated overlaps of these histotypes with the human counterparts, including clinical presentation, biologic behavior, tumor genetics, and treatment response. The molecular mechanisms that underlie canine BCL are still unknown and new studies to improve diagnosis, therapy, and the utilization of canine species as spontaneous animal tumor models are undeniably needed. Recent work using human DLBCL transcriptomes has suggested that long non-coding RNAs (lncRNAs) play a key role in lymphoma pathogenesis and pinpointed a restricted number of lncRNAs as potential targets for further studies. Results: To expand the knowledge of non-coding molecules involved in canine BCL, we used transcriptomes obtained from a cohort of 62 dogs with newly-diagnosed multicentric DLBCL, MZL and FL that had undergone complete staging work-up and were treated with chemotherapy or chemo-immunotherapy. We developed a customized R pipeline performing a transcriptome assembly by multiple algorithms to uncover novel lncRNAs, and delineate genome-wide expression of unannotated and annotated lncRNAs. Our pipeline also included a new package for high performance system biology analysis, which detects high-scoring network biological neighborhoods to identify functional modules. Moreover, our customized pipeline quantified the expression of novel and annotated lncRNAs, allowing us to subtype DLBCLs into two main groups. The DLBCL subtypes showed statistically different survivals, indicating the potential use of lncRNAs as prognostic biomarkers in future studies. Conclusions: In this manuscript, we describe the methodology used to identify lncRNAs that differentiate B-cell lymphoma subtypes and we interpreted the biological and clinical values of the results. We inferred the potential functions of lncRNAs to obtain a comprehensive and integrative insight that highlights their impact in this neoplasm

Development and validation of a gene expression oligo microarray for the gilthead sea bream (Sparus aurata)

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

fshr, a fish sex-determining locus shows variable incomplete penetrance across flathead grey mullet populations

Whole-genome sequence data were produced from a single flathead grey mullet female and assembled into a draft genome sequence, whereas publicly available sequence data were used to obtain a male draft sequence. Two pools, each consisting of 60 unrelated individuals respectively of male and female fish were analysed using Pool-Sequencing. Mapping and analysis of Pool-Seq data against the draft genome(s) revealed &gt;30 loci potentially associated with sex, the most promising locus of which, encoding the follicle stimulating hormone receptor (fshr) and harbouring two missense variants, was genotyped on 245 fish from four Mediterranean populations. Genotype data showed that fshr represents a previously unknown sex-determining locus, though the incomplete association pattern between fshr genotype and sex-phenotype, the variability of such pattern across different populations, and the presence of other candidate loci, reveal that a greater complexity underlies sex determination in the flathead grey mullet

New molecular and therapeutic insights into canine diffuse large B cell lymphoma elucidates the role of the dog as a model for human disease

open21siopenAresu, Luca; Ferraresso, Serena; Marconato, Laura; Cascione, Luciano; Napoli, Sara; Gaudio, Eugenio; Kwee, Ivo; Tarantelli, Chiara; Testa, Andrea; Maniaci, Chiara; Ciulli, Alessio; Hillmann, Petra; Bohnacker, Thomas; Wymann, Matthias P; Comazzi, Stefano; Milan, Massimo; Riondato, Fulvio; Dalla Rovere, Giulia; Giantin, Mery; Giannuzzi, Diana; Bertoni, FrancescoAresu, Luca; Ferraresso, Serena; Marconato, Laura; Cascione, Luciano; Napoli, Sara; Gaudio, Eugenio; Kwee, Ivo; Tarantelli, Chiara; Testa, Andrea; Maniaci, Chiara; Ciulli, Alessio; Hillmann, Petra; Bohnacker, Thomas; Wymann, Matthias P; Comazzi, Stefano; Milan, Massimo; Riondato, Fulvio; Dalla Rovere, Giulia; Giantin, Mery; Giannuzzi, Diana; Bertoni, Francesc