Vitellogenesis in the Patagonian toothfish (Dissostichus eleginoides) conditioned to a recirculating aquaculture system.

The Patagonian toothfish (Dissostichus eleginoides) is a new promising fish species for diversifying the aquaculture industry in Chile because of its high economic value and high international demand. However, when attempting to start aquaculture of a new species, one of the major challenges is successfully achieving conditions to reproduce them. This is particularly difficult when the information on the biology and physiology of the reproduction process of the species in question is scarce, as is the case with D.

Novel microsatellite markers discovery in Patagonian toothfish (Dissostichus eleginoides) using high-throughput sequencing.

Patagonian toohfish (Dissostichus eleginoides), is a sub Antartic notothenioid fish key in the marine ecosystem that sustains fishery of higher commercial value in the world. However, there are a scarce knowledge or information about its population genetic background, product of the almost null information of molecular markers available for this species. Here, we use high-throughput sequencing technology (Illumina platform) to develop 1071 microsatellite loci, of which 22 loci were selected to evaluation.

High density lipoproteins down-regulate transcriptional expression of pro-inflammatory factors and oxidative burst in head kidney leukocytes from rainbow trout, Oncorhynchus mykiss.

Teleosts are the first group of vertebrates possessing an acquired immune system; however, it is less developed than in mammals and is highly influenced by environmental changes. Therefore, innate immunity effectors play a more critical role in survival of pathogen-challenged fish. In a previous study we showed that trout high density lipoprotein (HDL), and its major apolipoprotein (ApoA-I) are widely expressed in primary defense barriers and other immune-relevant tissues, displaying important antibacterial activity in vitro.

Detection of up-regulated serum amyloid A transcript and of amyloid AA aggregates in skeletal muscle lesions of rainbow trout infected with Flavobacterium psychrophilum.

Serum amyloid A (SAA) is a family of acute-phase proteins, recognized as important effectors of innate immunity in higher vertebrates. Under pro-inflammatory conditions, up-regulation of saa transcripts occurs not only in the liver, but also in several extrahepatic tissues of a wide variety of vertebrates. SAA is also known as the precursor to amyloid A (AA), a major component of amyloid fibrils deposited in liver, kidney and spleen of humans suffering chronic inflammatory diseases.

Serum amyloid A: a typical acute-phase reactant in rainbow trout?

Acute serum amyloid A (A-SAA) has been considered a major acute-phase reactant and an effector of innate immunity in all vertebrates. The work presented here shows that the expression of A-SAA is strongly induced in a wide variety of immune-relevant tissues in rainbow trout, either naturally infected with Flavobacterium psychrophilum or challenged with lipopolysaccharide (LPS) or CpG oligonucleotides (CpG ODN). Nevertheless, A-SAA was undetectable by Western blot either in the plasma or in high-density lipoprotein (HDL) of infected or challenged fish, using either an anti-mouse SAA1 IgG or an anti-trout A-SAA peptide serum, which recognise both the intact recombinant trout A-SAA and fragments derived from it.

Apolipoprotein A-I, an antimicrobial protein in Oncorhynchus mykiss: evaluation of its expression in primary defence barriers and plasma levels in sick and healthy fish.

Antimicrobial proteins and peptides play an important role in the primary defence barriers in vertebrates and invertebrates. In a previous study it was shown that high-density lipoprotein (HDL) and its major apolipoproteins, ApoA-I and ApoA-II display antimicrobial activity in the carp (Cyprinus carpio L.).

Hexose transporters GLUT1 and GLUT3 are colocalized with hexokinase I in caveolae microdomains of rat spermatogenic cells.

Postmeiotic spermatogenic cells, but not meiotic spermatogenic cells respond differentially with glucose-induced changes in [Ca2+]i indicating a differential transport of glucose via facilitative hexose transporters (GLUTs) specifically distributed in the plasma membrane. Several studies have indicated that plasma membrane in mammalian cells is not homogeneously organized, but contains specific microdomains known as detergent-resistant membrane domains (DRMDs), lipid rafts or caveolae. The association of these domains and GLUTs isoforms has not been characterized in spermatogenic cells.

Undetectable apolipoprotein A-I gene expression suggests an unusual mechanism of dietary lipid mobilisation in the intestine of Cyprinus carpio.

High density lipoprotein (HDL) has been shown to play an important role in the dietary lipid mobilisation in the carp. In spite of this, previous studies have failed to demonstrate the synthesis of the major protein component of HDL, apolipoprotein A-I (apoA-I), in the proximal intestine of the carp. Therefore, the aim of the present study was to evaluate the expression of apoA-I throughout the entire intestine.

Apolipoproteins A-I and A-II are potentially important effectors of innate immunity in the teleost fish Cyprinus carpio.

We have previously shown that high density lipoprotein is the most abundant protein in the carp plasma and displays bactericidal activity in vitro. Therefore the aim of this study was to analyze the contribution of its principal apolipoproteins, apoA-I and apoA-II, in defense. Both apolipoproteins were isolated by a two step procedure involving affinity and gel filtration chromatography and were shown to display bactericidal and/or bacteriostatic activity in the micromolar range against Gram-positive and Gram-negative bacteria, including some fish pathogens.

Local expression of apolipoprotein A-I gene and a possible role for HDL in primary defence in the carp skin.

Antimicrobial proteins and peptides play an important role in the primary defence of epithelial barriers in vertebrates and invertebrates. Here we report the detection of the apolipoproteins A-I and A-II in the epidermis and epidermal mucus of the carp (Cyprinus carpio L.) by immunohistochemistry and Western blot analysis.

Specific binding of the endocytosis tracer horseradish peroxidase to intestinal fatty acid-binding protein (I-FABP) in apical membranes of carp enterocytes.

In a previous study we had demonstrated that a 15-kDa protein present in carp intestinal brush-border membrane vesicles (BBMV) was able to bind the endocytosis tracer horseradish peroxidase (HRP) with high specificity. Here we show that this protein corresponds to a peripheral membrane protein, identified by partial amino acid sequence analysis as the intestinal fatty acid-binding protein (I-FABP), a member of the small cytosolic fatty acid binding protein family (FABPs). The presence of I-FABP and its HRP-binding activity was demonstrated both in the cytosolic and membrane-associated fractions of intestinal mucosa by Western and ligand blot analyses, respectively.

Expression of GM-CSF receptors in male germ cells and their role in signaling for increased glucose and vitamin C transport.

We studied the expression and function of the granulocyte-macrophage colony stimulating factor (GM-CSF) receptor in male germ cells. RT-PCR showed expression of mRNAs encoding the alpha- and beta-subunits of the GM-CSF receptor in human testis, and the presence of the alpha- and beta-proteins was confirmed by immunoblotting with anti-alpha and anti-beta-antibodies. Immunolocalization studies showed the level of expression of GM-CSF alpha- and beta-subunits in the germ line in the testis and in ejaculated spermatozoa.

Horseradish peroxidase binding to intestinal brush-border membranes of Cyprinus carpio. Identification of a putative receptor.

Morphologic studies have shown that the classic endocytosis tracer horseradish peroxidase (HRP) is actively internalized by vesicular transport in the carp intestine, suggesting the existence of specific binding sites in the apical membrane of enterocytes. The aim of the present study was to develop an in vitro binding assay using isolated carp intestinal brush-border membranes (BBM) to demonstrate and characterize these specific HRP binding sites. The results obtained show that HRP binding to BBM exhibits a saturable mode and high affinity (K(d) = 22 nM).

Oral administration of insulin in winter-acclimatized carp (Cyprinus carpio) induces hepatic ultrastructural changes.

1. The intestinal absorption of insulin in carps was assessed examining the transepithelial passage of ingested gold-labeled hormone by electron microscopy. Insulin transfer occurred mainly through the intercellular spaces between the enterocytes.

Evidence that the putative COOH-terminal signal transamidase involved in glycosylphosphatidylinositol protein synthesis is present in the endoplasmic reticulum.

Nascent proteins destined to be processed to a glycosylphosphatidylinositol (GPI)-anchored membrane form contain NH2-terminal and COOH-terminal signal peptides. The first directs a nascent protein into the endoplasmic reticulum; the second peptide targets the protein to a putative COOH-terminal signal transamidase where cleavage of the peptide and addition of the GPI anchor occur. We recently showed that ATP hydrolysis is required for maturation of GPI proteins at a stage prior to transamidation.

Biosynthesis of glycosylphosphatidylinositol (GPI)-anchored membrane proteins in intact cells: specific amino acid requirements adjacent to the site of cleavage and GPI attachment.

Mutational studies were previously carried out at the omega site intact cells (Micanovic, R., L. Gerber, J.

Placental alkaline phosphatase: a model for studying COOH-terminal processing of phosphatidylinositol-glycan-anchored membrane proteins.

Placental alkaline phosphatase (PLAP) has been used as a model for studying the biosynthesis of the phosphatidylinositol-glycan (PI-G)-protein linkage in intact cells and in cell-free systems. However, for the study of processing in cell-free systems, a small protein devoid of glycosylation sites is preferable. A PLAP-derived cDNA was engineered that codes for a nascent protein (mini-PLAP) of 28 kDa in which the NH2- and COOH-termini are retained but most of the interior of PLAP is deleted.

Effect of seasonal acclimatization on estrogen-induced vitellogenesis and on the hepatic estrogen receptors in the male carp.

The levels of circulating vitellogenin in the plasma of male carp after induction by estradiol-17 beta was examined in summer- and winter-acclimatized fish. During the warm season male fish exhibited a clear vitellogenic response, whereas the fish adapted to the cold season did not. The evaluation of the hepatic estrogen receptors revealed that although steroid-binding affinity is not affected by seasonal acclimatization, the concentration of estrogen receptors certainly is.

Phosphatidylinositol-glycan (PI-G)-anchored membrane proteins: requirement of ATP and GTP for translation-independent COOH-terminal processing.

Placental alkaline phosphatase (PLAP) belongs to a class of proteins that are anchored to the plasma membrane by a COOH-terminal phosphatidylinositol-glycan (PI-G) moiety. Nascent forms of such proteins undergo NH2- and COOH-terminal processing to yield the mature PI-G-tailed proteins. We previously introduced a shortened engineered form of preproPLAP (preprominiPLAP) that permits monitoring in cell-free preparations its sequential processing to the pro form and then to the mature PI-G-tailed form.

Biosynthesis of phosphatidylinositol-glycan (PI-G)-anchored membrane proteins in cell-free systems: PI-G is an obligatory cosubstrate for COOH-terminal processing of nascent proteins.

It is generally recognized that nascent proteins destined to be processed to a phosphatidylinositol-glycan (PI-G)-anchored membrane form contain a hydrophobic signal peptide at both their NH2 and COOH termini. In previous studies we showed that rough microsomal membranes (RM) prepared from CHO cells can carry out COOH-terminal processing. We have now investigated RM prepared from many additional cell types, including frog oocytes, B cells, and T cells, and found that all are competent with respect to COOH-terminal processing.

Biosynthesis of phosphatidylinositol-glycan (PI-G)-anchored membrane proteins in cell-free systems: cleavage of the nascent protein and addition of the PI-G moiety depend on the size of the COOH-terminal signal peptide.

Nascent translation products of PI-G-anchored membrane proteins contain both NH2- and COOH-terminal signal sequences of approximately 15-30 residues that are removed during processing. Removal of the latter occurs concomitant with the addition of the PI-G moiety to the newly formed COOH terminus. In human placental alkaline phosphatase (PLAP) the COOH-terminal signal peptide contains 29 residues.

Cell-free processing of nascent proteins destined to be linked to the plasma membrane by a phosphatidylinositol-glycan anchor.

Certain proteins are anchored to the outer plasma membrane by a phosphatidylinositol-glycan (PI-G) linker. Nascent forms of PI-G anchored proteins contain both NH2- and COOH-terminal signal peptides. The function and structural requirements of the COOH-terminal signal peptide as discussed and some studies on the cell-free processing of a nascent protein to its mature PI-G tailed form are presented.

Interaction of cibacron blue and anilinonaphthalenesulphonate with lipoproteins provides a new means for simple isolation of these plasma proteins.

The selective interaction of serum proteins with immobilized Cibacron Blue and the binding properties of the dye anilinonaphthalenesulphonate has been used to separate albumin and lipoproteins by affinity chromatography. The novel binding of anilinonaphthalenesulphonate to lipoproteins from the sera of lamprey, fish and mammals provides a simple procedure for the isolation of these plasma proteins, and permit preparation of specific antisera, tools particularly relevant for evolutionary and clinical studies.