Author: Andrew Scholefield
Scholefield, Andrew, 2016 Fatty acid metabolism in southern bluefin tuna (Thunnus maccoyii), Flinders University, School of Biological Sciences
This electronic version is made publicly available by Flinders University in accordance with its open access policy for student theses. Copyright in this thesis remains with the author. This thesis may incorporate third party material which has been used by the author pursuant to Fair Dealing exceptions. If you are the owner of any included third party copyright material and/or you believe that any material has been made available without permission of the copyright owner please contact email@example.com with the details.
Southern bluefin tuna (SBT, Thunnus maccoyii) is a highly valuable aquaculture species but little is known of its exact nutritional requirements due to the prohibitively high costs of classical feeding trials with this species. As an alternative approach, this thesis reports on the use of a recently established SBT cell line (designated SBT-E1) to investigate fatty acid metabolism and mitochondrial biogenesis in this species. Addition of the C18 polyunsaturated fatty acids (PUFA) 18:3n-3 or 18:2n-6 to the cell culture medium had little effect on cell proliferation, whereas addition of the long-chain PUFA 20:4n-6, 20:5n-3 or 22:6n-3 significantly reduced cell proliferation, especially at higher concentrations and especially for DHA. Addition of vitamin E to the culture medium overcame this effect, suggesting that it was due to oxidative stress. The fatty acid profiles of the total lipid from the cells reflected those of the respective culture media except that 22:6n-3 was substantially more abundant in the cells than in the media. Fatty acid esterification occurred predominantly into phosphatidylcholine and phosphatidylethanolamine, the two most abundant lipid classes. The SBT-E1 cells showed very limited Δ6 fatty acyl desaturase (Fads) activity towards either 18:3n-3 or 18:2n-6 but substantial elongation of very long chain fatty acids (Elovl) activity towards 20:5n-3. The latter activity is usually attributable to an Elovl5 enzyme. Surprisingly though, there were much higher levels of Δ6Fads compared with Elovl5 gene expression in the SBT-E1 cells, suggesting that a different Elovl enzyme may catalyse this reaction in SBT. The cells also showed substantial β-oxidation of 18:3n-3 and 20:5n-3 but much less activity towards 18:0, 18:1n-9 or 18:2n-6. These results may explain the high 22:6n-3 to 20:5n-3 ratios found in the SBT tissue lipids. Serum deprivation did not significantly affect gene expression of the mitochondrial markers citrate synthase (CS) and cytochrome c oxidase subunit 1 (COX1) in the SBT-E1 cells but it did significantly upregulate the expression of peroxisome proliferator activator receptor γ (PPARγ) co-activator 1 α (PGC-1α). This may indicate a role for PGC-1α in the cellular response to reduced fatty acid availability. Interestingly, the cells supplied with the reduced concentration of serum together with 20:5n-3 did not exhibit significantly increased PGC-1α gene expression, indicating that 20:5n-3 was able to overcome some of the effects of the reduced serum concentration. To further investigate the role of PGC-1α in SBT, a PGC-1α cDNA was cloned from SBT liver. This is the first report of a PGC-1α sequence from any tuna species. The PGC-1α cDNA corresponded to what is recognised as the canonical PGC-1α. When comparing different SBT tissues, PGC-1α gene expression was found to be high in the red muscle and the expression levels in the different tissues correlated well with the expression of its co-activation target, PPARγ. In contrast, the expression of PGC-1α did not appear to influence mitochondrial abundance, as indicated by the gene expression of CS and COX1. The results are discussed in light of the unusual physiology of tunas.
Keywords: Fatty acids, Metabolism, Tuna, Polyunsaturated fatty acids, Docosahexanoic acid, Eicosapentanoic acid
Subject: Biological Sciences thesis
Thesis type: Doctor of Philosophy
School: School of Biological Sciences
Supervisor: Kathryn Schuller