Project description:A greater understanding of the proteins involved in reproduction can benefit animal production. New advances in proteomics are having a major impact on our understanding of how spermatozoa acquire their capacity for fertilization [1]. Sperm proteomics aims at the identification of the proteins that compose the sperm cell and the study of their function [2]. The sperm cell is one of the most highly differentiated cells and is composed of a head with a highly compacted chromatin structure and a large flagellum with midpiece that contains the required machinery for movement and therefore to deliver the paternal genetic and epigenetic content to the oocyte [3]. By being so highly differentiated, spermatozoa are advantageous cells to study proteomics of specific compartments such as the membrane, which basically is the area of major importance for its role in interacting with the surroundings and the oocyte [4]. The fusion of a sperm and an oocyte is a sophisticated process that must be preceded by suitable changes in the sperm's membrane composition [5]. Recent studies of spermatozoa from the proteomic point of view have allowed the identification of different proteins in spermatozoa that are responsible for the regulation of normal/defective sperm functions [6]. While several techniques are available in proteomics, LC-MS based analysis of complex protein/peptide mixtures has turned out to be a mainstream analytical technique for quantitative proteomics [7]. Using this method, detailed proteomic data are now available for human [8], macaque [9,10], mouse [11], rat [12], bull [13-15], stallion [16], fruit fly [17], Caenorhabditis elegans [18], carp [19], rainbow trout [20], mussel [21], ram [22], honeybee [23] and rooster [24] sperm membrane proteins. Rabbit (Oryctolagus cuniculus) is an important mammalian species worldwide, being at the same time of commercial interest and a research model animal. European rabbit meat production is approximately 500 thousand tons, corresponding to a 30% share of world production [25]. Besides, rabbits account for the seventh highest number of animals slaughtered per year in the European Union-27, with 347,603 × 1000 head in 2014 [26]. In a previous work, we identified and quantified rabbit seminal plasma proteins between two different genotypes [27], concluding the clear effect of genotype in the abundance of certain seminal plasma proteins. However, it is unknown at present whether these differences also exist at sperm proteome level. Therefore, the aim of the present study was to characterise rabbit sperm membrane proteins through NanoLC-MS/MS analysis focusing on the influence of the genetic origin.
Project description:Although IgD first appeared, along with IgM, in the cartilaginous fishes and has been retained throughout subsequent vertebrate evolution, it has been lost in a diverse group of vertebrate species. We previously showed that, unlike vertebrates that express IgD, the rabbit lacks an IgD (Cδ) gene within 13.5 kb downstream of the IgM gene. We report here that, by conducting BLAST searches of rabbit Ig heavy chain genomic DNA with known mammalian IgD exons, we identified the remnant of the rabbit Cδ gene approximately 21 kb downstream of the IgM gene. The remnant Cδ locus lacks the δCH1 and hinge exons, but contains truncated δCH2 and δCH3 exons, as well as largely intact, but non-functional, secretory and transmembrane exons. In addition, we report that the Cδ gene probably became non-functional in leporids at least prior to the divergence of rabbits and hares ~12 million years ago.
Project description:BackgroundStem cells are capable of unlimited self-renewal and are able to remain undifferentiated for extended periods of time prior to their differentiation into specific cell lineages. Because of the issues (ethical and religious) involved in the use of embryonic stem cells and the limited plasticity of adult stem cells, an alternative cell source could be foetal stem cells derived from extra-embryonic tissue, which are highly proliferative, grow in vitro and possess interesting immunogenic characteristics. As a result, the amniotic membrane of several species has been studied as an important new source of stem cells.MethodsHere, we cultured and characterized mesenchymal progenitor cells derived from the rabbit amniotic membrane, and investigated their differentiation potential. In total, amniotic membranes were collected from eight rabbit foetuses and were isolated by the explant technique. The obtained cells were cultured in DMEM-HIGH glucose and incubated at 37 °C in a humidified atmosphere with 5% CO2.ResultsThe cells adhered to the culture plates and showed a high proliferative capacity with fibroblast-like morphologies. The cells showed a positive response for markers for the cytoskeleton, mesenchymal stem cells and proliferation, pluripotency and haematopoietic precursor stem cells. However, the cells were negative for CD45, a marker of haematopoietic cells. Furthermore, the cells had the capacity to be induced to differentiate into osteogenic, adipogenic and chondrogenic lineages. In addition, when the cells were injected into nude mice, we did not observe the formation of tumours.ConclusionsIn summary, our results demonstrate that multipotent mesenchymal stem cells can be obtained from the rabbit amniotic membrane for possible use in future cell therapy applications.
Project description:Although the rabbit genome has already been annotated, it is mobilome remains largely unknown. Here, multiple pipelines were used to de novo mine and annotate the mobilome in rabbit. Four families and 19 subfamilies of LINE1s, two families and nine subfamilies of SINEs, and 12 ERV families were defined in rabbit based on sequence identity, structural organization, and phylogenetic tree. The analysis of insertion age and polymerase chain reaction suggests that a number of families are very young and may remain active, such as L1B, L1D, OcuSINEA, and OcuERV1. RepeatMasker annotation revealed a distinct transposable element landscape within the genome, with approximately two million copies of SINEs, representing the greatest proportion of the genome (19.61%), followed by LINEs (15.44%), and LTRs (4.11%), respectively, considerably different from most other mammal mobilomes except hedgehog and tree shrew, in which LINEs have the highest proportion. Furthermore, a very high rate of insertion polymorphisms (>85%) for the youngest subfamily (OcuSINEA1) was identified by polymerase chain reaction. The majority of retrotransposon insertions overlapped with protein-coding regions (>80%) and lncRNA (90%) genes. Genomic distribution bias was observed for retrotransposons, with those immediately upstream (-1 kb) and downstream (1 kb) of genes significantly depleted. Local GC content in 50-kb widows had significantly negative correlations with LINE (rs=-0.996) and LTR (rs=-0.829) insertions. The current study revealed a distinct mobilome landscape in rabbit, which will assist in the elucidation of the evolution of the genome of lagomorphs, and even other mammals.
Project description:BackgroundFur is an important genetically-determined characteristic of domestic rabbits; rabbit furs are of great economic value. We used the Solexa sequencing technology to assess gene expression in skin tissues from full-sib Rex rabbits of different phenotypes in order to explore the molecular mechanisms associated with fur determination.Methodology/principal findingsTranscriptome analysis included de novo assembly, gene function identification, and gene function classification and enrichment. We obtained 74,032,912 and 71,126,891 short reads of 100 nt, which were assembled into 377,618 unique sequences by Trinity strategy (N50=680 nt). Based on BLAST results with known proteins, 50,228 sequences were identified at a cut-off E-value ≥ 10-5. Using Blast to Gene Ontology (GO), Clusters of Orthologous Groups (KOG) and Kyoto Encyclopedia of Genes and Genomes (KEGG), we obtained several genes with important protein functions. A total of 308 differentially expressed genes were obtained by transcriptome analysis of plaice and un-plaice phenotype animals; 209 additional differentially expressed genes were not found in any database. These genes included 49 that were only expressed in plaice skin rabbits. The novel genes may play important roles during skin growth and development. In addition, 99 known differentially expressed genes were assigned to PI3K-Akt signaling, focal adhesion, and ECM-receptor interactin, among others. Growth factors play a role in skin growth and development by regulating these signaling pathways. We confirmed the altered expression levels of seven target genes by qRT-PCR. And chosen a key gene for SNP to found the differentially between plaice and un-plaice phenotypes rabbit.Conclusions/significanceThe rabbit transcriptome profiling data provide new insights in understanding the molecular mechanisms underlying rabbit skin growth and development.