Long-chain polyunsaturated fatty acid synthesis in fish: Comparative analysis of Atlantic salmon (Salmo salar L.) and Atlantic cod (Gadus morhua L.) Delta 6 fatty acyl desaturase gene promoters

Fish vary in ability to biosynthesise n-3 long-chain polyunsaturated fatty acids (LC-PUFA), with marine fish such as cod being inefficient in comparison to freshwater and salmonid fish. We investigated differences in the gene promoters of Δ6 fatty acyl desaturase (Δ6 FAD), a critical enzyme in LC-PUFA biosynthesis, in cod and salmon. Progressive deletions and targeted mutations of the promoters were tested for activity in a transfected fish cell line under low or high LC-PUFA treatment, and regions sufficient to direct transcription were identified. Comparison of these regions with sequences of corresponding regions of Δ6 FAD genes from mammals, amphibians and fish indicated a remarkable conservation of binding sites for SREBPs and NF-Y. In addition to these sites, a site was identified in salmon with similarity to that recognised by Sp1 transcription factor, and which was required for full expression of the salmon Δ6 FAD gene. The cod promoter was less active and lacked the Sp1 site. Eicosapentaenoic acid suppressed LC-PUFA synthesis in AS cells and also suppressed activity of the salmon Δ6 FAD promoter although this activity was likely mediated through sites other than Sp1, possibly similar to those recognised by NF-Y and SREBP transcription factors

Effect of partial substitution of dietary fish oil by vegetable oils on desaturation and beta-oxidation of [1-14C]18:3n-3 (LNA) and [1-14C]20:5n-3 (EPA) in hepatocytes and enterocytes of European sea bass (Dicentrarchus labrax L.)

The increasing worldwide aquaculture output and concomitant decrease in the stocks of feed-grade fish used for fish oil production has made fish oil replacement in feeds a priority for the aquaculture industry. The regulation of fatty acid metabolism in fish is important in order to determine strategies for the best use of plant oils in diets for commercially important cultured fish species. We have studied the desaturation/elongation and β-oxidation of 14C-linolenic (LNA) and 14C-eicosapentaenoic (EPA) acids in hepatocytes and pyloric caecal enterocytes in European sea bass fed diets with partial substitution (60%) of fish oil (FO) with vegetable oils (rapeseed, linseed and palm oil) blended in different proportions, for 64 weeks. The rate of desaturation of 14C‑LNA was very low in hepatocytes from all treatments and no significant differences were observed among treatments. The rate of desaturation of 14C‑LNA in enterocytes was higher than that in hepatocytes but still low (less than 5% of total radioactivity recovered). The desaturation of 14C‑EPA in enterocytes was also higher than in hepatocytes, but again was low and no significant differences were found among treatments. The rates of ß‑oxidation of 14C-LNA and 14C-EPA were much higher than the rates of desaturation in both hepatocytes and enterocytes, however, no significant differences were observed in either hepatocytes or enterocytes among treatments. The rates of β-oxidation of 14C-LNA were considerably higher than those of 14C-EPA in both hepatocytes and enterocytes. In conclusion, European sea bass (a carnivorous marine fish), showed very low desaturation and elongation of LNA to EPA and DHA, and EPA to DHA, higher β−oxidation of LNA than EPA, and all desaturation and oxidation activities were significantly higher in enterocytes than in hepatocytes. A second major conclusion is that no clear quantitative nutritional effects on the desaturation/elongation and β−oxidation activities in either hepatocytes or enterocytes of sea bass were observed upon the inclusion of vegetable oils in the diet

Alien Registration- Tocher, Christina (Baldwin, Cumberland County)

https://digitalmaine.com/alien_docs/32859/thumbnail.jp

GM crop ban: how Scottish salmon – and public health – could have benefited from this technology

First paragraph: The plan to ban the growing of genetically modified crops is disappointing to many scientists. It would be highly unsatisfactory if, as it appears, such an important decision has been made by the Scottish government without a proper informed debate that takes the scientific evidence fully into consideration. It is not enough for the rural affairs secretary, Richard Lochhead, to say that he is not prepared to “gamble” with the future of Scotland’s £14bn food and drink sector.https://theconversation.com/gm-crop-ban-how-scottish-salmon-and-public-health-could-have-benefited-from-this-technology-4598

Environmental and dietary influences on highly unsaturated fatty acid biosynthesis and expression of fatty acyl desaturase and elongase genes in liver of Atlantic salmon (Salmo salar)

Highly unsaturated fatty acid (HUFA) synthesis in Atlantic salmon (Salmo salar) was known to be influenced by both nutritional and environmental factors. Here we aimed to test the hypothesis that both these effectors involved similar molecular mechanisms. Thus, HUFA biosynthetic activity and the expression of fatty acyl desaturase and elongase genes were determined at various points during an entire two year production cycle in salmon fed diets containing either 100% fish oil or diets in which a high proportion (75% and 100%) of fish oil was replaced by C18 polyunsaturated fatty acid-rich vegetable oil. The results showed that HUFA biosynthesis in Atlantic salmon varied during the growth cycle with peak activity around seawater transfer and subsequent low activities in seawater. Consistent with this, gene expression of Δ6 desaturase, the rate-limiting step in the HUFA biosynthetic pathway, was highest around the point of seawater transfer and lowest during the seawater phase. In addition, the expression of both Δ6 and Δ5 desaturase genes was generally higher in fish fed the vegetable oil-substituted diets compared to fish fed fish oil, particularly in the seawater phase. Again, generally consistent with this, the activity of the HUFA biosynthetic pathway was invariably higher in fish fed diets in which fish oil was substituted by vegetable oil compared to fish fed only fish oil. In conclusion, these studies showed that both nutritional and environmental modulation of HUFA biosynthesis in Atlantic salmon involved regulation of fatty acid desaturase gene expression

Alien Registration- Tocher, William (Baldwin, Cumberland County)

https://digitalmaine.com/alien_docs/32860/thumbnail.jp

Metabolism and functions of lipids and fatty acids in teleost fish

Lipids and their constituent fatty acids are, along with proteins, the major organic constituents of fish, and they play major roles as sources of metabolic energy for growth including reproduction, and movement including migration. Furthermore, the fatty acids of fish lipids are rich in ω3 long chain, highly unsaturated fatty acids (n-3 HUFA) that have particularly important roles in animal nutrition, including fish and human nutrition, reflecting their roles in critical physiological processes. Indeed, fish are the most important food source of these vital nutrients for man Thus, the long standing interest in fish lipids stems from their abundance and their uniqueness. This review attempts to summarise our present state of knowledge of various aspects of the basic biochemistry, metabolism and functions of fatty acids, and the lipids they constitute part of, in fish, seeking where possible to relate that understanding as much to fish in their natural environment as to farmed fish. In doing so, it highlights the areas that require to be investigated in greater depth and also the increasing application of molecular technologies in fish lipid metabolism which will fascilitate further advances through molecular biological and genetic techniques including genomics and proteomics

Cytosolic cholesterol ester hydrolase in adrenal cortex

Cholesterol ester hydrolase (CEH) in adrenocortical cytosol was known to be phosphorylated and activated, in response to ACTH in a cAMPdependent protein kinase mediated process. The purification of CEH from bovine adrenocortical cytosol was attempted. The use of detergents to solubilise the enzyme from lipid-rich aggregates was investigated and sodium cholate was found to be effective. A purification procedure using cholate solubilised enzyme was developed. The detergent interfered in the operation of the ion-exchange columns, and pure enzyme was not obtained. Hydrophobic chromatography was studied but it was found to be unsuitable for the purification of CEH under the conditions investigated. An affinity chromatography technique was developed using columns of glass beads coated non-covalently with cholesterol oleate. Some positive results were obtained but the low capacity of the columns combined with the low concentration of the enzyme in the tissue cytosol prevented further study of the purified activity. A further purification procedure utilising a non-ionic detergent and gradient sievorptive chromatography resulted in a 150-fold purification of CEH from bovine adrenocortical cytosol with a recovery of about 25%. The specific activity of the partially purified enzyme (CEHQ2 preparation) was approximately 60 nmol oleic acid produced.min 3.mg protein Labelling studies using the CEHQ2 preparation, [γ-32P] ATP and [1,3-3H] DFP suggested that the enzyme activity was associated with a protein with Mr approximately 84 000. A phosphoprotein phosphatase with Mr 35 000 wTas purified from bovine adrenocortical cytosol to a state approaching homogeneity. The purified 32 phosphatase was active when measured towards P-phosphoprotexn and p-nitrophenyl phosphate. The role of this phosphoprotein phosphatase in the modulation of cytosolic CEH activity was investigated, but the enzyme did not deactivate partially-purified CEH from bovine adrenocortical cytosol. Cytosolic CEH in rat adrenal was found to exhibit a diurnal variation in activity. The enzyme activity was significantly higher during the dark phase. Serum corticosterone concentration reflected this variation in CEH activity. In vivo experiments including acute ACTH administration and dexamethasone suppression of pituitary ACTH secretion suggested that ACTH was involved in the production and maintenance of the diurnal rhythm of rat adrenal cytosolic CEH activity. The protein components of bovine adrenocortical and rat adrenal lipid droplets were investigated. The lipid droplets from both species contained a major protein subunit with Mr 40 000 and several other minor proteins. The protein profile was similar in both species. The Mr 40 000 apoprotein was able to be phosphorylated in rat lipid droplets but not in bovine lipid droplets. The delipidated apoprotein was phosphorylated in both cases. Bovine adrenocortical lipid droplets contained about 4% by weight protein, but CEH was found to be only a minor component of the protein fraction

Omega-3 long-chain polyunsaturated fatty acids and aquaculture in perspective

In the 40 years since the essentiality of polyunsaturated fatty acids (PUFA) in fish was first established by determining quantitative requirements for 18:3n−3 and 18:2n−6 in rainbow trout, essential fatty acid (EFA) research has gone through distinct phases. For 20 years the focus was primarily on determining qualitative and quantitative EFA requirements of fish species. Nutritional and biochemical studies showed major differences between fish species based on whether C18 PUFA or long-chain (LC)-PUFAwere required to satisfy requirements. In contrast, in the last 20 years, research emphasis shifted to determining “optimal” levels of EFA to support growth of fish fed diets with increased lipid content and where growth expectations were much higher. This required greater knowledge of the roles and functions of EFA inmetabolismand physiology, and howthese impacted on fish health and disease. Requirement studies were more focused on early life stages, in particular larval marine fish, defining not only levels, but also balances between different EFAs. Finally, a major driver in the last 10–15 years has been the unavoidable replacement of fish oil and fishmeal in feeds and the impacts that this can have on n−3 LC-PUFA contents of diets and farmed fish, and the human consumer. Thus, dietary n−3 infish feeds can be defined by three levels. Firstly, the minimumlevel required to satisfy EFA requirements and thus prevent nutritional pathologies. This level is relatively small and easy to supply even with today's current high demand for fish oil. The second level is that required to sustainmaximumgrowth and optimumhealth in fish being fed modern high-energy diets. The balance between different PUFA and LC-PUFA is important and defining them is more challenging, and so ideal levels and balances are still not well understood, particularly in relation to fish health. The third level is currently driving much research; how can we supply sufficient n−3 LC-PUFA tomaintain these nutrients in farmed fish at similar or higher levels than in wild fish? This level far exceeds the biological requirements of the fish itself and to satisfy it we require entirely new sources of n−3 LCPUFA. We cannot rely on the finite and limited marine resources that we can sustainably harvest or efficiently recycle. We need to produce n−3 LC-PUFA de novo and all possible options should be considered