New Insights on Intermediary Metabolism for a Better Understanding of Nutrition in Teleosts.

The rapid development of aquaculture production throughout the world over the past few decades has led to the emergence of new scientific challenges to improve fish nutrition. The diet formulations used for farmed fish have been largely modified in the past few years. However, bottlenecks still exist in being able to suppress totally marine resources (fish meal and fish oil) in diets without negatively affecting growth performance and flesh quality. A better understanding of fish metabolism and its regulation by nutrients is thus mandatory. In this review, we discuss four fields of research that are highly important for improving fish nutrition in the future: ( a) fish genome complexity and subsequent consequences for metabolism, ( b) microRNAs (miRNAs) as new actors in regulation of fish metabolism, ( c) the role of autophagy in regulation of fish metabolism, and ( d) the nutritional programming of metabolism linked to the early life of fish.

Effect of food shortage and temperature on age 0+ salmonids: a contribution to predict the effects of climate change.

Brown trout Salmo trutta alevins were maintained at 8 and 11° C at three conditions over a 9 day period from yolk sac exhaustion: fed ad libitum, starved or fed ad libitum after starvation. Whole-body gene expressions for proteins involved in energy metabolism and the two primary proteolytic pathways were assessed. This study is the first to show an over-expression of proteasome and autophagy-related genes in young stages of salmonids, particularly at 11° C.

Myostatin and the skeletal muscle atrophy and hypertrophy signaling pathways.

Myostatin, a member of the transforming growth factor-ß superfamily, is a potent negative regulator of skeletal muscle growth and is conserved in many species, from rodents to humans. Myostatin inactivation can induce skeletal muscle hypertrophy, while its overexpression or systemic administration causes muscle atrophy. As it represents a potential target for stimulating muscle growth and/or preventing muscle wasting, myostatin regulation and functions in the control of muscle mass have been extensively studied. A wealth of data strongly suggests that alterations in skeletal muscle mass are associated with dysregulation in myostatin expression. Moreover, myostatin plays a central role in integrating/mediating anabolic and catabolic responses. Myostatin negatively regulates the activity of the Akt pathway, which promotes protein synthesis, and increases the activity of the ubiquitin-proteasome system to induce atrophy. Several new studies have brought new information on how myostatin may affect both ribosomal biogenesis and translation efficiency of specific mRNA subclasses. In addition, although myostatin has been identified as a modulator of the major catabolic pathways, including the ubiquitin-proteasome and the autophagy-lysosome systems, the underlying mechanisms are only partially understood. The goal of this review is to highlight outstanding questions about myostatin-mediated regulation of the anabolic and catabolic signaling pathways in skeletal muscle. Particular emphasis has been placed on (1) the cross-regulation between myostatin, the growth-promoting pathways and the proteolytic systems; (2) how myostatin inhibition leads to muscle hypertrophy; and (3) the regulation of translation by myostatin.

Leucine limitation regulates myf5 and myoD expression and inhibits myoblast differentiation.

Satellite cells are the major pool of muscle stem cells after birth; they represent an important component required to maintain muscle mass and functionality during life. The molecular mechanisms involved in myogenic differentiation are relatively well-known. However, the role of extracellular stimulus in the control of differentiation remains largely unresolved. Notably little is known about the impact of nutrients on this process. Here we have studied the role of leucine, an essential amino acid, in the control of myogenic differentiation. Leucine is a well-known regulator of muscle protein synthesis. It acts not only as a substrate for translation but also as a regulator of gene expression and signaling pathways such as those involving mTOR and GCN2. In this study we demonstrated that the lack of leucine abolishes the differentiation of both C2C12 myoblasts and primary satellite cells. This effect is associated with a modification of the pattern of expression of the myogenic regulatory factors (MRF) myf5 and myoD. We report an up-regulation of myf5 mRNA and a decrease of myoD protein level during leucine starvation. This study demonstrates the importance of a nutrient, leucine, in the control of the myogenic differentiation program.

Metformin improves postprandial glucose homeostasis in rainbow trout fed dietary carbohydrates: a link with the induction of hepatic lipogenic capacities?

Carnivorous fish are poor users of dietary carbohydrates and are considered to be glucose intolerant. In this context, we have tested, for the first time in rainbow trout, metformin, a common anti-diabetic drug, known to modify muscle and liver metabolism and to control hyperglycemia in mammals. In the present study, juvenile trout were fed with very high levels of carbohydrates (30% of the diet) for this species during 10 days followed by feeding with pellets supplemented with metformin (0.25% of the diet) for three additional days. Dietary metformin led to a significant reduction in postprandial glycemia in trout, demonstrating unambiguously the hypoglycemic effect of this drug. No effect of metformin was detected on mRNA levels for glucose transporter type 4 (GLUT4), or enzymes involved in glycolysis, mitochondrial energy metabolism, or on glycogen level in the white muscle. Expected inhibition of hepatic gluconeogenic (glucose-6-phosphatase, fructose-1,6-bisphosphatase, and phosphoenolpyruvate carboxykinase) mRNA levels was not found, showing instead paradoxically higher mRNA levels for these genes after drug treatment. Finally, metformin treatment was associated with higher mRNA levels and activities for lipogenic enzymes (fatty acid synthase and glucose-6-phosphate dehydrogenase). Overall, this study strongly supports that the induction of hepatic lipogenesis by dietary glucose may permit a more efficient control of postprandial glycemia in carnivorous fish fed with high carbohydrate diets.

Characterization and comparison of fatty acyl Delta6 desaturase cDNAs from freshwater and marine teleost fish species.

Fish are the most important dietary source of the n-3 highly unsaturated fatty acids (HUFA), eicosapentaenoic (EPA) and docosahexaenoic acid (DHA), that have particularly important roles in human nutrition reflecting their roles in critical physiological processes. The objective of the study described here was to clone, functionally characterize and compare expressed fatty acid desaturase genes involved in the production of EPA and DHA in freshwater and marine teleost fish species. Putative fatty acid desaturase cDNAs were isolated and cloned from common carp (Cyprinus carpio) and turbot (Psetta maximus). The enzymic activities of the products of these cDNAs, together with those of cDNAs previously cloned from rainbow trout (Oncorhynchus mykiss) and gilthead sea bream (Sparus aurata), were determined by heterologous expression in the yeast Saccharomyces cerevisiae. The carp and turbot desaturase cDNAs included open reading frames (ORFs) of 1335 and 1338 base pairs, respectively, specifying proteins of 444 and 445 amino acids. The protein sequences possessed all the characteristic features of microsomal fatty acid desaturases, including three histidine boxes, two transmembrane regions, and N-terminal cytochrome b(5) domains containing the haem-binding motif, HPGG. Functional expression showed all four fish cDNAs encode basically unifunctional Delta6 fatty acid desaturase enzymes responsible for the first and rate-limiting step in the biosynthesis of HUFA from 18:3n-3 and 18:2n-6. All the fish desaturases were more active towards the n-3 substrate with 59.5%, 31.5%, 23.1% and 7.0% of 18:3n-3 being converted to 18:4n-3 in the case of turbot, trout, sea bream and carp, respectively. The enzymes also showed very low, probably physiologically insignificant, levels of Delta5 desaturase activity, but none of the products showed Delta4 desaturase activity. The cloning and characterization of desaturases from these fish is an important advance, as they are species in which there is a relative wealth of data on the nutritional regulation of fatty acid desaturation and HUFA synthesis, and between which substantive differences occur.

Cloning and nutritional regulation of a Delta6-desaturase-like enzyme in the marine teleost gilthead seabream (Sparus aurata).

Marine fish are presumed to have a lower capacity than freshwater fish for the bioconvertion of 18C fatty acids into 20-22C highly unsaturated fatty acids (HUFA). The present work investigated the first step of this pathway, the Delta6-desaturation, in gilthead seabream. A full-length desaturase-like cDNA was identified from total RNA extracted from viscera of juvenile fish fed for 96 days on an experimental HUFA-free diet containing olive oil as the sole lipid source. The open reading frame encodes a 445-amino acid peptide that contains two membrane-spanning domains, three histidine-rich regions, and a cytochrome b(5) domain, which are characteristic of Delta6- and Delta5-desaturases. Predicted protein sequence of seabream desaturase-like indicated a high percentage of identity with mammalian Delta6-desaturases (approx. 65%). Northern analysis showed two transcripts of approximately 3.7 and 1.8 kb which were highly expressed in fish fed on HUFA-free diet and slightly expressed in fish fed on HUFA-rich diet. The fatty acid profile of the former group was characterized by high levels of Delta6-desaturation products (18:2 n-9 and 20:2 n-9) with no detectable levels of Delta5-desaturation product (20:3n-9). These results demonstrate for the first time the presence and nutritional modulation of a Delta6-desaturase-like cDNA in a marine fish.

Cloning, tissue distribution and nutritional regulation of a Delta6-desaturase-like enzyme in rainbow trout.

This report describes the cloning, nutritional regulation and tissue distribution of a desaturase-like enzyme in rainbow trout (Oncorhynchus mykiss). The open reading frame of the trout desaturase-like cDNA encodes a 454-amino acid peptide that contains two membrane-spanning domains, three histidine-rich regions and a cytochrome b5 domain, which all align perfectly with the same domains located in other recently identified vertebrate Delta5- and Delta6-desaturases. Nutritional regulation of trout desaturase-like gene expression, as well as the tissue expression profile, are also similar to those observed in other vertebrate Delta5- and Delta6-desaturases. Finally, the sequence alignments between the predicted protein sequence of rainbow trout desaturase-like and other Delta6- and Delta5-desaturases revealed a high percentage identity with Delta6-desaturases (64-66% identity with vertebrate Delta6-desaturases). These results demonstrate for the first time the presence and nutritional modulation of a Delta6-desaturase-like cDNA in rainbow trout.

Regulation of expression of the rat SOCS-3 gene in hepatocytes by growth hormone, interleukin-6 and glucocorticoids mRNA analysis and promoter characterization.

Suppressors of cytokine signalling (SOCS) represent a newly discovered family of molecules that seem to play an important role in the shutting off of cytokine and possibly peptide hormone action. Thus, understanding the mechanisms controlling their expression is of cardinal importance. In the present study, we have cloned the rat SOCS-3 gene and analyzed its expression and the functioning of its promoter in hepatocytes. Expression of SOCS-3 mRNA, which is very weak in freshly isolated cells, tended to increase when hepatocytes were incubated without hormones. Growth hormone (GH) and, to a much larger extent, interleukin-6 (IL-6) rapidly activated mRNA synthesis whereas glucocorticoids (GC) strongly inhibited both basal and hormone-dependent expressions. A short promoter fragment (-137/+35) responded maximally to GH and IL-6 (a threefold stimulation for each effector) and to GC (a 70-80% inhibition), whereas longer promoter sequences supported higher basal activity and lower positive hormonal responses. Deletion and mutation analyses indicated that all hormonal responses were dependent on two cis-acting sequences termed the G-rich and the A/T-rich elements. Only the A/T-rich element was active in a heterologous context, thus behaving as a typical enhancer. Unexpectedly, the two signal transducer and activator of transcription (STAT) binding sites found immediately upstream of the G-rich motif didn't seem to participate in either GH or IL-6 effect, despite the fact that one of them strongly responded to IL-6 when placed in front of a heterologous promoter. Finally, the negative regulation of SOCS-3 promoter by GC that may contribute to gene silencing in vivo, appeared to involve interactions of the GC receptor with other transcription factors and not direct binding to DNA, as no GC-response element was found in the sequence.

Definition of an amino-terminal domain of the human T-cell leukemia virus type 1 envelope surface unit that extends the fusogenic range of an ecotropic murine leukemia virus.

Murine leukemia viruses (MuLV) and human T-cell leukemia viruses (HTLV) are phylogenetically highly divergent retroviruses with distinct envelope fusion properties. The MuLV envelope glycoprotein surface unit (SU) comprises a receptor-binding domain followed by a proline-rich region which modulates envelope conformational changes and fusogenicity. In contrast, the receptor-binding domain and SU organization of HTLV are undefined. Here, we describe an HTLV/MuLV envelope chimera in which the receptor-binding domain and proline-rich region of the ecotropic MuLV were replaced with the potentially corresponding domains of the HTLV-1 SU. This chimeric HTLV/MuLV envelope was processed, specifically interfered with HTLV-1 envelope-mediated fusion, and similar to MuLV envelopes, required cleavage of its cytoplasmic tail to exert significant fusogenic properties. Furthermore, the HTLV domain defined here broadened ecotropic MuLV envelope-induced fusion to human and simian cell lines.