A description of the origins, design and performance of the TRAITS-SGP Atlantic salmon Salmo salar L. cDNA microarray.

The origins, design, fabrication and performance of an Atlantic salmon microarray are described. The microarray comprises 16 950 Atlantic salmon-derived cDNA features, printed in duplicate and mostly sourced from pre-existing expressed sequence tag (EST) collections [SALGENE and salmon genome project (SGP)] but also supplemented with cDNAs from suppression subtractive hybridization libraries and candidate genes involved in immune response, protein catabolism, lipid metabolism and the parr-smolt transformation. A preliminary analysis of a dietary lipid experiment identified a number of genes known to be involved in lipid metabolism. Significant fold change differences (as low as 1.2x) were apparent from the microarray analysis and were confirmed by quantitative real-time polymerase chain reaction analysis. The study also highlighted the potential for obtaining artefactual expression patterns as a result of cross-hybridization of similar transcripts. Examination of the robustness and sensitivity of the experimental design employed demonstrated the greater importance of biological replication over technical (dye flip) replication for identification of a limited number of key genes in the studied system. The TRAITS (TRanscriptome Analysis of Important Traits of Salmon)-salmon genome project microarray has been proven, in a number of studies, to be a powerful tool for the study of key traits of Atlantic salmon biology. It is now available for use by researchers in the wider scientific community.

Interferon type I and type II responses in an Atlantic salmon (Salmo salar) SHK-1 cell line by the salmon TRAITS/SGP microarray.

Interferons (IFNs) are cytokines that have proinflammatory, antiviral, and immunomodulatory effects and play a central role during a host response to pathogens. The IFN family contains both type I and type II molecules. While there are a number of type I IFNs, there is only one type II IFN. Recently both type I and type II IFN genes have been cloned in salmonid fish and recombinant proteins produced showing IFN activity. We have stimulated an Atlantic salmon cell line (SHK-1) with both type I and type II recombinant salmonid IFNs and analyzed the transcriptional response by microarray analysis. Cells were exposed to recombinant IFNs for 6 or 24 h or left unexposed as controls. RNA was hybridized to an Atlantic salmon cDNA microarray (salmon 17K feature TRAITS/SGP array) in order to assess differential gene expression in response to IFN exposure. For IFN I and II, 47 and 72 genes were stimulated, respectively; most genes were stimulated by a single IFN type, but some were affected by both IFNs, indicating coregulation of the IFN response in fish. Real-time PCR analysis was employed to confirm the microarray results for selected differentially expressed genes in both a cell line and primary leukocyte cultures.

Feed efficiency of rainbow trout can be improved through selection: different genetic potential on alternative diets.

To assess the genetic potential for selection of increased feed efficiency in rainbow trout (Oncorhynchus mykiss), we estimated the heritabilities and correlations for BW, daily weight gain (DG), and daily feed intake (DFI). Body weight was recorded 5 times, and DG and DFI 3 times during a feeding trial lasting 22 mo. To test the hypothesis that phenotypic and genetic parameters were influenced by a nutritional environment, fish were fed either a modern normal protein diet (NP, 40 to 45% protein and 30 to 33% lipid) or an alternative high protein diet (HP, 50 to 56% protein, 20 to 24% lipid) in a split-family design. Results showed that there were no large differences in heritabilities between the diets. Average heritability for DFI over both diets and different fish ages was low (average h2 = 0.10), indicating that modest genetic changes in response to selection can be obtained. Average heritabilities for BW and DG over both diets and different fish ages were 0.28 and 0.33, respectively. The NP diet enabled fish to express a wide range of BW, as shown by the increased coefficients of phenotypic variation for BW. Fish fed the HP diet showed increased phenotypic variation for DFI in > 750-g fish. On the NP diet, genetic correlations of DFI with DG and BW were very strong for 750- to 2,000-g fish. In contrast, on the HP diet, the respective correlations were moderate to low, revealing more genetic potential to change growth and feed intake simultaneously in opposite directions. An analysis of the predicted selection responses showed that selection solely for high DG improved feed efficiency as a correlated genetic response. Simultaneous selection for high DG and reduced DFI, in turn, may increase genetic gain in feed efficiency by a factor of 1.2 compared with selection solely for DG. However, variation for growth and feed intake and the relationships between these traits were different in different nutritional environments, leading to divergent genetic responses on the alternative diets.

Transcriptome response following administration of a live bacterial vaccine in Atlantic salmon (Salmo salar).

Antibacterial responses have been studied in Atlantic salmon following an acute intra peritoneal injection of a genetically attenuated (aroA(-)) strain of Aeromonas salmonicida known to elicit protective immunity. Three tissues were studied for transcriptional changes, the liver, head kidney and the gill. RNA was collected from fish 6, 12, 24 and 48 h following infection or at the same time points from fish injected with PBS as non-infected control. PCR-select cDNA subtraction libraries were constructed from pooled 24 and 48 h post infection RNA to identify up-regulated mRNAs. One thousand four hundred and eighty six cDNA clones were sequenced from enriched cDNA libraries, of which 71% had significant homologies to known functional proteins. Many of these clones have previously been un-characterised in Atlantic salmon. A salmonid cDNA microarray was used to further analyse the gene expression profile as the library construction in itself does not answer the dynamics of the response. The greatest increase in expression identified in the array analysis was a liver antibacterial peptide, hepcidin that was increased 11-fold following the challenge. A panel of clones were chosen for semiquantitative reverse transcriptase PCR from all time points sampled. These results indicated there were both temporal differences and tissue differences in the transcriptional response to bacterial exposure, potentially of relevance to the establishment of protection.

Protein growth rate in rainbow trout (Oncorhynchus mykiss) is negatively correlated to liver 20S proteasome activity.

The efficiency with which fish and other animals add and maintain body proteins is a balance between synthesis of proteins and their degradation. In fish that have similar food consumption and protein synthesis rates, a greater ratio of synthesis to degradation would be expected to produce more efficient conversion of food into growth. In addition, we hypothesised that high activities of the proteasome, a major pathway of protein degradation, would be negatively correlated with growth rate. In order to test this hypothesis we maintained rainbow trout for 62 days, during which repeat measurements of food consumption and growth were made. We selected fish for high and low growth efficiencies. Protein degradation was estimated from the difference between protein synthesis (determined by 15N flux) and protein growth. We found that protein synthesis rates were significantly higher in the low growth efficiency group, as were estimated protein degradation rates. In another group of fish that also did not differ in food consumption, the activity of the proteasome in the liver, but not in the muscle, was negatively correlated with growth rates. These two experiments showed that high proteasome activity is linked to decreased growth efficiency.

Proteomic sensitivity to dietary manipulations in rainbow trout.

Changes in dietary protein sources due to substitution of fish meal by other protein sources can have metabolic consequences in farmed fish. A proteomics approach was used to study the protein profiles of livers of rainbow trout that have been fed two diets containing different proportions of plant ingredients. Both diets control (C) and soy (S) contained fish meal and plant ingredients and synthetic amino acids, but diet S had a greater proportion of soybean meal. A feeding trial was performed for 12 weeks at the end of which, growth and protein metabolism parameters were measured. Protein growth rates were not different in fish fed different diets; however, protein consumption and protein synthesis rates were higher in the fish fed the diet S. Fish fed diet S had lower efficiency of retention of synthesised protein. Ammonia excretion was increased as well as the activities of hepatic glutamate dehydrogenase and aspartate amino transferase (ASAT). No differences were found in free amino acid pools in either liver or muscle between diets. Protein extraction followed by high-resolution two-dimensional electrophoresis, coupled with gel image analysis, allowed identification and expression of hundreds of protein. Individual proteins of interest were then subjected to further analysis leading to protein identification by trypsin digest fingerprinting. During this study, approximately 800 liver proteins were analysed for expression pattern, of which 33 were found to be differentially expressed between diets C and S. Seventeen proteins were positively identified after database searching. Proteins were identified from diverse metabolic pathways, demonstrating the complex nature of gene expression responses to dietary manipulation revealed by proteomic characterisation.

Ubiquitin-proteasome-dependent proteolysis in rainbow trout (Oncorhynchus mykiss): effect of food deprivation.

The ubiquitin-proteasome proteolytic pathway is a major route of protein degradation and of particular importance in muscle proteolysis in mammals. In this study, the beta proteasome subunit N3 and polyubiquitin genes of the rainbow trout, Oncorhynchus mykiss, were sequenced and tissue distribution of gene expression was examined. The effects of 14-day food withdrawal were assessed on the N3 subunit and polyubiquitin gene expression in terms of mRNA, 20S proteasome proteolytic activity and ubiquitin protein abundance in trout liver and muscle. Both sequences are highly conserved, and the rainbow trout ubiquitin amino acid sequence is identical to the mammalian protein. The proteasome beta subunit N3 has 92% similarity to the Xenopus sequence. Starvation halved the polyubiquitin mRNA level in liver but had no effect on muscle levels. No significant effect of food withdrawal was observed on the proteasome mRNA in liver or muscle. Food withdrawal decreased the 20S proteasome proteolytic activity and the abundance of ubiquitin protein in both muscle and liver. Co-regulation of the proteasome and ubiquitin was indicated by the high correlation ( R=0.924) between 20S activity and ubiquitin abundance. Overall, this study demonstrates that starvation down-regulates the ubiquitin-proteasome pathway, possibly highlighting differences in the regulation of protein turnover in poikilothermic and endothermic animals.

Protein synthesis costs could account for the tissue-specific effects of sub-lethal copper on protein synthesis in rainbow trout (Oncorhynchus mykiss).

This study investigates protein synthesis, following exposure to sub-lethal Cu, in rainbow trout in vivo and in vitro. The investigation has two aims: to determine if perturbations in protein synthesis, compared with other physiological changes, are a biomarker of Cu pollution and to evaluate the most productive role of cellular models in ecotoxicology. Protein synthesis rates were measured by labelling with 3H-phenylalanine. In vivo this was applied by a single (i.p.) injection and in vitro by bathing the cells in 3H-phenylalanine labelled culture media. The effects in vivo were tissue specific. After 3 weeks' exposure to 0.7 microM Cu only skin protein synthesis was reduced. Gills and liver from the same fish were unaffected. This reduction in skin protein synthesis appears to be more sensitive than some other biomarkers reported in the literature. However, Cu concentrations greater by orders of magnitude were required to reproduce this reduction in protein synthesis in skin cell explants (200 and 400 microM). Hepatocyte protein synthesis was unaffected by 10, 20 and 40 microM Cu and a separate investigation has also shown that 25 and 75 microM Cu does not effect protein synthesis in cultured gill cells. Oxygen consumption rates were also measured in vitro by monitoring the decline in O2 partial pressure. The Cu concentrations given above resulted in a decline in O2 consumption rates in the respective cell types. By measuring protein synthesis and O2 consumption after treatment with a protein synthesis inhibitor (cycloheximide), the costs of protein synthesis were also determined. Synthesis costs in hepatocytes are close to the theoretical minimum and are only marginally affected by Cu. Gill cell synthesis costs are also minimal and are unaffected. In skin explants, the reduction in protein synthesis was accompanied by greatly increased synthesis costs. This in vitro result offers a hypothesis as to the tissue-specific effects in vivo; i.e. the energetic demand of protein synthesis may determine tissue sensitivity or susceptibility. Cell or tissue types with high protein synthesis rates are able to avoid detrimental increases in the synthesis cost when exposed to Cu. In tissues with a low protein synthesis rate any further reduction is more likely to incur a potentially damaging increase in protein synthesis costs. Thus, whilst in vitro models may have little practical use in environmental monitoring, they may be best used as a mechanistic tool in understanding susceptibility or tolerance to sub-lethal Cu.

Growth, feeding frequency, protein turnover, and amino acid metabolism in European lobster Homarus gammarus L.

The effect of feeding frequency on growth and protein metabolism in the European lobster, Homarus gammarus, was investigated. Fourth (IV) stage lobsters H. gammarus were fed individually a marine animal meal (herring/mussels meal) for 56 days. Feeding a daily ration equivalent to 10% of their body weight gave better growth than feeding daily rations of 5% and 20%. Protein synthesis rates were similar for the three food rations but protein growth rates were significantly lower and protein degradation rates highest in the 5% body weight per day ration group. The efficiency with which synthesised protein was retained as growth was found to be 38% in the in the 10% ratio group. Protein synthesis rates of lobsters were found to be lower than those for shrimps (Penaeus vannamei). The amino acid flux also suggests a lower protein conversion efficiency than shrimps P. vannamei. The results suggests that lobsters are slow, periodic feeders and that growth can be readily increased by manipulation of particular environmental factors such as feeding frequency.

Complete suppression of protein synthesis during anoxia with no post-anoxia protein synthesis debt in the red-eared slider turtle Trachemys scripta elegans.

Two previous studies of the effects of anoxia on protein synthesis in anoxia-tolerant turtles (Trachemys scripta elegans, Chrysemys picta bellii) have generated opposing results. Using the flooding-dose method, we measured the rate of protein synthesis following injection and incorporation of a large dose of radiolabelled phenylalanine to resolve the question of whether anoxia results in a downregulation of protein synthesis. After 1 h of anoxia, levels of protein-incorporated radiolabel indicated that protein synthesis rates in the intestine, heart, liver, brain, muscle and lungs were not significantly different from those of normoxic controls. However, from 1 to 6 h of anoxia, quantities of protein-incorporated radiolabel did not increase, suggesting that protein synthesis had ceased or had decreased below a measurable level. There was also no significant post-anoxia increase in protein synthesis rates above normoxic control levels during 3 h of recovery from anoxia. RNA-to-protein ratios did not change significantly in any tissue except the heart, in which RNA levels decreased below normoxic control levels after 6 h of anoxia. Except in the heart, downregulation of protein synthesis during anoxia does not appear to be mediated by changes in tissue RNA concentration.

RNA turnover and protein synthesis in fish cells.

Protein synthesis in fish has been previously correlated with RNA content. The present study investigates whether protein and RNA synthesis rates are similarly related. Protein and RNA synthesis rates were determined from 3H-phenylalanine and 3H-uridine incorporation, respectively, and expressed as % x day(-1) and half-lives, respectively. Three fibroblast cell lines were used: BF-2, RTP, CHSE 214, which are derived from the bluegill, rainbow trout and Chinook salmon, respectively. These cells contained similar RNA concentrations (approximately 175 microg RNA x mg(-1) cell protein). Therefore differences in protein synthesis rates, BF-2 (31.3 +/- 1.8)>RTP (25.1 +/- 1.7)>CHSE 214 (17.6 +/-1.1), were attributable to RNA translational efficiency. The most translationally efficient RNA (BF-2 cells), 1.8 mg protein synthesised x microg(-1) RNA x day(-1), corresponded to the lowest RNA half-life, 75.4 +/- 6.4 h. Translationally efficient RNA was also energetically efficient with BF-2 cells exploiting the least costly route of nucleotide supply (i.e. exogenous salvage) 3.5-6.0 times more than the least translationally efficient RNA (CHSE 214 cells). These data suggest that differential nucleotide supply, between intracellular synthesis and exogenous salvage, constitutes the area of pre-translational flexibility exploited to maintain RNA synthesis as a fixed energetic cost component of protein synthesis.

Tissue-specific changes in RNA synthesis in vivo during anoxia in crucian carp.

The overall energy budget for protein synthesis (i.e., transcription plus translation) is thought to consist of fixed and variable components, with RNA synthesis accounting for the former and protein synthesis the latter. During anoxia, the downregulation of protein synthesis (i.e., the variable component), to reduce energetic demand, is an important aspect of survival in crucian carp. The present study examines RNA synthesis during anoxia by labeling with [(3)H]uridine. A novel synthesis rate calculation is presented, which allows for the tissue-specific salvage of uridine, with synthesis rates finally expressed relative to DNA. After 48 h anoxia, the decline (29%) in brain RNA synthesis and increases in the heart and liver (132 and 871%, respectively) support known RNA functions during hypoxic/anoxic survival. This study provides evidence that, in an anoxia-tolerant species, survival mechanisms involving RNA are able to operate because tissue-specific restructuring of the RNA synthesis process enables fixed synthesis costs to be maintained; this may be as vital to survival as exploiting the variable energetic demand of protein synthesis.

Changes in physiological parameters and feeding behaviour of Atlantic salmon Salmo salar infected with sea lice Lepeophtheirus salmonis.

Atlantic salmon Salmo salar L. artificially infected with salmon lice Lepeophtheirus salmonis (Krøyer 1837) recovered from detrimental physiological changes and skin damage induced by preadult lice as the parasites matured. Growth rates of Atlantic salmon remained unaffected by lice infection, but food consumption decreased with increasing feeding and movement of the lice prior to and post-mating, correlating with the appearance of head erosions and detrimental changes in physiological integrity. Food consumption of the fish increased as the lice moulted to the adult stage and gravid female lice settled in a posterior location on the fish, subsequently reducing the impact of infection and allowing recovery of the skin damage. However, the impact of preadults was limited, as the decrease in food consumption of fish at 21 d post-infection had no effect on either the specific growth rate or condition factor of the fish. Furthermore, the intensity of lice infections at each of the sample days was not correlated with food consumption, specific growth rate or any of the haematological or physiological parameters measured, either before or after infection, indicating that lice intensity was independent of social dominance/subordinance. This work has provided the first evidence that infected fish can recover from the detrimental changes caused by lice infection, even when they are still infected with lice. If fish can survive the preadult stage of lice, then the mortal impact of lice infections is greatly reduced.

Effects of salmon lice Lepeophtheirus salmonis on sea trout Salmo trutta at different times after seawater transfer.

The physiological and behavioural effects and skin damage caused by salmon lice Lepeophtheirus salmonis (Krøyer) infections on sea trout Salmo trutta L. smolts were greater in fish infected with lice 2 wk after seawater transfer than in fish infected 6 wk after seawater transfer. The initial prevalence of infection was 100% for both groups and the intensity of infection decreased significantly with time over 5 wk. Significantly fewer of the fish infected 2 wk after seawater transfer had resumed feeding by the end of the experiment, leading to a loss of body condition. Furthermore, these fish suffered more severe damage to the skin and detrimental changes in physiological integrity than fish infected 6 wk after seawater transfer as a direct consequence of feeding preadult lice, leading to osmoregulatory failure and death. Although this study was carried out in laboratory conditions, results indicate that lice infections may potentially have a detrimental impact on the survival of wild smolts after seawater transfer.

Gill protein turnover: costs of adaptation.

Measurements of gill protein synthesis, and hence turnover, were greatly facilitated over the last decade by the application of "flooding dose" methodology to non-mammalian species. Numerous studies show that in fish and aquatic invertebrates, gills are among the most active tissues with respect to protein turnover, this being true under a variety of environmental and nutritional conditions. The main components being turned over in fish gills are probably collagen, primarily in the gill arches, and epithelial cell proteins in the filaments, both arches and filaments having similar protein synthesis rates. Intriguingly, differences are apparent between protein synthesis rates of adjacent holobranchs, the first (most anterior) being significantly more active than the second or third, perhaps hinting at functional differences between holobranchs. Experimental estimates of energetic costs for protein synthesis, derived from cycloheximide treatment of isolated perfused gills, give a maximum value of 14 mmol O2/g protein synthesized, which is about double theoretical costs. Environmental stressors, such as heavy metals or acid/aluminum, have variable effects on branchial protein turnover. Limited data suggest that zinc or acid exposure depresses protein synthesis, whereas acid/aluminum increases it quite markedly. Calculations indicate that whereas effects within the gills may be substantial, in terms of whole animal energetics, the costs of branchial adaptation are likely to be small.

Estradiol stimulates growth hormone production in female goldfish.

The effects of estradiol (E2) on growth hormone (GH) production was investigated in gonad-intact female goldfish. It was first necessary to generate a specific antibody for use in immunocytochemistry, Western, and dot-blot analyses of GH production. To accomplish this, grass carp GH (gcGH) cDNA was cloned by the reverse transcription polymerase chain reaction (RT-PCR) and expressed in Echerichia coli and a specific polyclonal antibody to recombinant gcGH was generated in the rabbit. In Western blot, the anti-gcGH antibody specifically immunoreacted with recombinant gcGH, purified natural common carp GH, and with a single 21.5-kDa GH form from pituitary extracts of grass carp, common carp, goldfish, and zebrafish but not salmon, trout, or tilapia. Intraperitoneal injection of the recombinant gcGH enhanced the growth rates of juvenile common carp demonstrating biological activity of this GH preparation. Electron microscopic studies showed that the anti-gcGH-I antibody specifically reacted with GH localized in the secretory granules of the goldfish somatotroph. Using anti-gcGH-I in a dot-blot assay, it was found that in vivo implantation of solid silastic pellets containing E2, (100 micrograms/g body weight for 5 days) increased pituitary GH content by 150% in female goldfish. In a second, independent study employing a previously characterized anticommon carp GH antibody for radioimmunoassay, it was found that E2 increased pituitary GH content by 170% and serum GH levels by approximately 350%. The E2-induced hypersecretion of GH and increase in pituitary GH levels was not associated with changes in steady-state pituitary GH mRNA levels, suggesting that this sex steroid may enhance GH synthesis at the posttranscriptional or translational level. Previous observations indicate that GH can stimulate ovarian E2 production. The present results show that E2 can in turn stimulate GH production, indicating the existence of a novel pituitary GH-ovarian feedback system in goldfish.

Tissue-specific changes in protein synthesis rates in vivo during anoxia in crucian carp.

Mechanisms of anoxia tolerance were investigated in crucian carp. Rates of protein synthesis were calculated in selected tissues of normoxic and anoxic animals. Exposure to 48 h of anoxia resulted in a significant reduction in protein synthesis in the liver (> 95%), heart (53%), and red and white muscle (52 and 56%, respectively), whereas brain protein synthesis rates were unaffected. Seven days of anoxia produced similar results. After 24 h of recovery from a 48-h anoxic period, protein synthesis rates had virtually returned to normoxic values. The effect of anoxia on the amount of RNA (relative to protein) varied depending on the tissue and also the length of exposure (except in the brain, where it was consistently reduced). However, the effect on RNA translational efficiency was purely tissue specific (i.e., independent of exposure time) and was unaffected in the heart, reduced in the liver and red and white muscle, and increased in the brain. Downregulation of protein synthesis on a tissue-specific basis appears to be a significant mechanism for energy conservation as well as maintaining neural function, thus promoting survival during anoxia.

Feed consumption, growth and growth efficiency of rainbow trout (Oncorhynchus mykiss (Walbaum)) fed on diets containing a bacterial single-cell protein.

The aim of the present study was to compare the nutritive value of bacterial single-cell protein (BSCP) with that of fishmeal in rainbow trout (Oncorhynchus mykiss (Walbaum)). Four diets were formulated to contain a total of 458 g crude protein/kg of which 0% was from BSCP in diet 1 (BSCP-0), 25% in diet 2 (BSCP-25), 62.5% in diet 3 (BSCP-62.5) and 100% in diet 4 (BSCP-100); the remainder of the protein was from fishmeal. There were two studies: in study 1, duplicate groups of twenty-five fish were fed on one of the four experimental diets at the rate of 20 g/kg body weight per d for 132 d. Feed consumption rates of individual fish were measured using radiography and the overall apparent absorption efficiency for N in each group was measured over a 2-week period. In study 2, N intake, consumption, absorption and accretion were measured for each fish under controlled environmental conditions (12 h: 12 h light-dark regime; 14 degrees). Higher dietary levels of BSCP resulted in significantly higher feed consumption rates but reduced N absorption efficiency and growth rates. However, a diet containing 25% BSCP (75% fishmeal) did not significantly influence growth rates, feed consumption and absorption efficiency compared with a 100% fishmeal diet. The N growth efficiencies were highest in fish fed on the diet containing the highest level of fishmeal and significantly decreased with increasing BSCP content. Construction of N budgets demonstrated that the reduction in growth in fish eating an increasingly larger proportion of BSCP was due to a decrease in N absorption and an increase in the excretion of urea.

Effect of feeding on the tissue free amino acid concentrations in rainbow trout (Oncorhynchus mykiss Walbaum).

This study investigates whether tissue free amino acid (FAA) pools in rainbow trout,Oncorhynchus mykiss (Walbaum), are altered following feeding and the relationships between the amount of food consumed and the FAA pool size. Trout were starved for 7 days to provide baseline data and then refed on day 8. Individual food intake was measured by radiography and the consumption of amino acids (AA) calculated from dietary protein consumption. Total FAA concentrations in the stomach, liver and white muscle were little changed at various times after the meal and this pattern was repeated for the majority of individual FAA. Overall, the most notable change was a reduction in essential FAA concentrations (principally in valine, leucine and isoleucine) in the white muscle following feeding. However, in the caeca total FAA, total essential FAA and a number of individual FAA were significantly elevated at 4, 9 and 15h following feeding. There were few significant correlations between dietary amino acid consumption and total tissue FAA and essential FAA concentration in the stomach, caecum and white muscle; correlations were stronger in the liver. In order to explain the relative constancy of total FAA concentrations in the tissues following food intake (which represents over 100% of the total FAA pool) a model is presented that quantifies the AA flux through the free pools and considers the role of protein turnover in regulating FAA pool size.

Protein synthesis, growth and energetics in larval herring (Clupea harengus) at different feeding regimes.

Rates of growth, protein synthesis and oxygen consumption were measured in herring larvae, Clupea harengus, in order to estimate the contribution that protein synthesis makes to oxygen consumption during rapid growth at 8°C. Protein synthesis rates were determined in larvae 9 to 17 d after hatching. Larvae were bathed in (3)H phenylalanine for several hours and the free pool and protein-bound phenylalanine specific radioactivities were determined.Fractional rates of protein synthesis increased 5 to 11 fold with feeding after a period of fasting. Efficiencies of retention of synthesized protein were approximately 50% during rapid growth. Rapid growth in herring larvae thus appears to be characterized by moderate levels of protein turnover similar to those obtained for larger fish. Increases in growth rate occurred without changes in RNA concentration, i.e., the larvae increased the efficiency of RNA rapidly. Oxygen consumption rates were not correlated with growth rates. Protein synthesis was estimated to account for 79% of the oxygen consumption, and energy costs of protein synthesis were high, i.e., about 98 mmole O2 g(-1) protein synthesized.