Data set for the proteomic inventory and quantitative analysis of chicken eggshell matrix proteins during the primary events of eggshell mineralization and the active growth phase of calcification

This research was funded by the French National Research Agency ANR (ANR-13-BSV6-0007-01, ANK-13-BSV6-0007-02 and ANK-13-BSV6-0007-05). The high resolution mass spectrometer was financed (SMHART project, 35069) by the European Regional Development Fund (ERDF), the Conseil Regional du Centre, the French National Institute for Agricultural Research (INRA) and the French National Institute of Health and Medical Research (Inserm), ARN acknowledges funding through Grants CGL2011-25906 (Ministerio de Economia, Spain).Chicken eggshell is a biomineral composed of 95% calcite calcium carbonate mineral and of 3.5% organic matrix proteins. The assembly of mineral and its structural organization is controlled by its organic matrix. In a recent study [1], we have used quantitative proteomic, bioinformatic and functional analyses to explore the distribution of 216 eggshell matrix proteins at four key stages of shell mineralization defined as: (1) widespread deposition of amorphous calcium carbonate (ACC), (2) ACC transformation into crystalline calcite aggregates, (3) formation of larger calcite crystal units and (4) rapid growth of calcite as columnar structure with preferential crystal orientation. The current article detailed the quantitative analysis performed at the four stages of shell mineralization to determine the proteins which are the most abundant. Additionally, we reported the enriched GO terms and described the presence of 35 antimicrobial proteins equally distributed at all stages to keep the egg free of bacteria and of 81 proteins, the function of which could not be ascribed.French National Research Agency (ANR) ANR-13-BSV6-0007-01 ANK-13-BSV6-0007-02 ANK-13-BSV6-0007-05European Union (EU) 35069Region Centre-Val de LoireFrench National Institute for Agricultural Research (INRA)Institut National de la Sante et de la Recherche Medicale (Inserm)Ministerio de Economia, Spain CGL2011-2590

Protein expression reveals a molecular sexual identity of avian primordial germ cells at pre-gonadal stages

International audienceIn poultry, in vitro propagated primordial germ cells (PGCs) represent an important tool for the cryopreservation of avian genetic resources. However, several studies have highlighted sexual differences exhibited by PGCs during in vitro propagation, which may compromise their reproductive capacities. To understand this phenomenon, we compared the proteome of pregonadal migratory male (ZZ) and female (ZW) chicken PGCs propagated in vitro by quantitative proteomic analysis using a GeLC-MS/MS strategy. Many proteins were found to be differentially abundant in chicken male and female PGCs indicating their early sexual identity. Many of the proteins more highly expressed in male PGCs were encoded by genes localised to the Z sex chromosome. This suggests that the known lack of dosage compensation of the transcription of Z-linked genes between sexes persists at the protein level in PGCs, and that this may be a key factor of their autonomous sex differentiation. We also found that globally, protein differences do not closely correlate with transcript differences indicating a selective translational mechanism in PGCs. Male and female PGC expressed protein sets were associated with differential biological processes and contained proteins known to be biologically relevant for male and female germ cell development, respectively. We also discovered that female PGCs have a higher capacity to uptake proteins from the cell culture medium than male PGCs. This study presents the first evidence of an early predetermined sex specific cell fate of chicken PGCs and their sexual molecular specificities which will enable the development of more precise sex-specific in vitro culture conditions for the preservation of avian genetic resources

ViSEAGO: Easier data mining of biological functions organized into clusters using Gene Ontology and semantic similarity

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ViSEAGO: a bioconductor package for clustering biological functions using gene ontology and semantic similarity

International audienceThe main objective of ViSEAGO package is to carry out a data mining of biological functions and establish links between genes involved in the study. We developed ViSEAGO in R to facilitate functional Gene Ontology (GO) analysis of complex experimental design with multiple comparisons of interest. It allows to study large-scale datasets together and visualize GO profiles to capture biological knowledge. The acronym stands for three major concepts of the analysis: Visualization, Semantic similarity and Enrichment Analysis of Gene Ontology. It provides access to the last current GO annotations, which are retrieved from one of NCBI EntrezGene, Ensembl or Uniprot databases for several species. Using available R packages and novel developments, ViSEAGO extends classical functional GO analysis to focus on functional coherence by aggregating closely related biological themes while studying multiple datasets at once. It provides both a synthetic and detailed view using interactive functionalities respecting the GO graph structure and ensuring functional coherence supplied by semantic similarity. ViSEAGO has been successfully applied on several datasets from different species with a variety of biological questions. Results can be easily shared between bioinformaticians and biologists, enhancing reporting capabilities while maintaining reproducibility. ViSEAGO is publicly available on

Identification of uterine ion transporters for mineralisation precursors of the avian eggshell

<p>Abstract</p> <p>Background</p> <p>In <it>Gallus gallus</it>, eggshell formation takes place daily in the hen uterus and requires large amounts of the ionic precursors for calcium carbonate (CaCO₃). Both elements (Ca²⁺, HCO₃^-) are supplied by the blood via trans-epithelial transport. Our aims were to identify genes coding for ion transporters that are upregulated in the uterine portion of the oviduct during eggshell calcification, compared to other tissues and other physiological states, and incorporate these proteins into a general model for mineral transfer across the tubular gland cells during eggshell formation.</p> <p>Results</p> <p>A total of 37 candidate ion transport genes were selected from our database of overexpressed uterine genes associated with eggshell calcification, and by analogy with mammalian transporters. Their uterine expression was compared by qRTPCR in the presence and absence of eggshell formation, and with relative expression levels in magnum (low Ca²⁺/HCO₃^-movement) and duodenum (high rates of Ca²⁺/HCO₃^- trans-epithelial transfer). We identified overexpression of eleven genes related to calcium movement: the TRPV6 Ca²⁺ channel (basolateral uptake of Ca²⁺), 28 kDa calbindin (intracellular Ca²⁺ buffering), the endoplasmic reticulum type 2 and 3 Ca²⁺ pumps (ER uptake), and the inositol trisphosphate receptors type 1, 2 and 3 (ER release). Ca²⁺ movement across the apical membrane likely involves membrane Ca²⁺ pumps and Ca²⁺/Na⁺ exchangers. Our data suggests that Na⁺ transport involved the SCNN1 channel and the Na⁺/Ca²⁺ exchangers SLC8A1, 3 for cell uptake, the Na⁺/K⁺ ATPase for cell output. K⁺ uptake resulted from the Na⁺/K⁺ ATPase, and its output from the K⁺ channels (KCNJ2, 15, 16 and KCNMA1).</p> <p>We propose that the HCO₃^- is mainly produced from CO₂ by the carbonic anhydrase 2 (CA2) and that HCO₃^- is secreted through the HCO₃^-/Cl^- exchanger SLC26A9. HCO₃^- synthesis and precipitation with Ca²⁺ produce two H⁺. Protons are absorbed via the membrane’s Ca²⁺ pumps ATP2B1, 2 in the apical membrane and the vacuolar (H+)-atpases at the basolateral level. Our model incorporate Cl^- ions which are absorbed by the HCO₃^-/Cl^- exchanger SLC26A9 and by Cl^- channels (CLCN2, CFTR) and might be extruded by Cl^-/H⁺ exchanger (CLCN5), but also by Na⁺ K⁺ 2 Cl^- and K⁺ Cl^- cotransporters.</p> <p>Conclusions</p> <p>Our <it>Gallus gallus</it> uterine model proposes a large list of ion transfer proteins supplying Ca²⁺ and HCO₃^-and maintaining cellular ionic homeostasis. This avian model should contribute towards understanding the mechanisms and regulation for ionic precursors of CaCO₃, and provide insight in other species where epithelia transport large amount of calcium or bicarbonate.</p

CHAPTER 14. Bioactive Minor Egg Components

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Identification par une approche innovante de protéomique à haut débit (diaPASEF) de protéines plasmatiques différentiellement exprimées au cours de la maturation sexuelle des truites Arc-en-ciel (Oncorhynchus mykiss)

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Cryopreservation effect on DNA methylation profile in rainbow trout spermatozoa

Abstract Spermatozoa are the cells that are most commonly used for cryopreservation of valuable genetic resources in aquaculture. It is known that fish spermatozoa transmit to the embryo not only their genetic but also their epigenetic profile, especially DNA methylation. Therefore, any alteration of the DNA methylation profile in spermatozoa induces the risk of transmitting epigenetic alterations to the offspring. The aim of this study was to assess the effect of cryopreservation on DNA methylation in rainbow trout spermatozoa. To trigger variable cellular response after freezing–thawing, spermatozoa from mature males were cryopreserved with dimethyl sulfoxide, methanol or glycerol as cryoprotectant. We observed that dimethyl sulfoxide was the best to preserve thawed spermatozoa functions. Methanol only slightly preserved all the cellular parameters, while glycerol failed to protect motility and fertilization ability. The consequences on DNA methylation were assessed using Reduced Representation Bisulfite Sequencing (RRBS). Sperm cryopreservation did not thoroughly impact DNA methylation, although 335–564 differentially methylated cytosines were characterized depending on the cryoprotectant. Very few of them were shared between cryoprotectants, and no correlation with the extent of cellular damage was found. Our study showed that DNA methylation was only slightly altered after sperm cryopreservation, and this may render further analysis of the risk for the progeny very challenging