Introduction
Polychaetes are promising sources of ω3 long-chain (≥C20) polyunsaturated fatty acids (LC-PUFA) due to their ability to synthesise these essential nutrients through the action of fatty acyl elongases (Elovl) , front-end desaturases (Fed), and methyl-end desaturases (ω-des) (Monroig et al., 2022). It has been suggested that s alinity can influence lipid metabolism in aquatic invertebrates , as it triggers osmotic stress that alters membrane lipid composition (Palacios and Racotta , 2007). Iron also plays a crucial role in lipid metabolism and, among other functions, iron is a cofactor of desaturase enzymes like Fed and ω-des involved in LC-PUFA biosynthesis (Shanklin et al., 2009). We hypothesize that salinity conditions leading to increased expression of Fed and/or ω-des will only result in increase desaturase activity and ultimately LC-PUFA biosynthesis, if iron supplementation ensures full activation of the desaturase enzyme pool. To test this hypothesis, this study investigated (i) the effects of salinity on fatty acid profiles and expression of genes involved in the ω3 LC-PUFA synthesis in the marine polychaete Platynereis dumerilii, and (ii) the combined effects of salinity and dietary iron on the P. dumerilii LC-PUFA biosynthesis.
Materials and Methods
Two in vivo trials were conducted using P. dumerilii juveniles maintained under standard conditions (18 °C, 16L:8D photoperiod ; low-PUFA diet ). Trial 1 assessed the effects of three salinity levels (30‰, 35‰, and 40‰) , using three plastic boxes per salinity level, each containing 16 worms. In Trial 2 , P. dumerilii were reared under four conditions combining two salinities (35‰ and 40‰) and two diets (with or without iron supplementation ), using four plastic boxes per condition, each with 25 worms . At the end of each trial, samples were collected to evaluate the effects on the ω3 LC-PUFA biosynthesis through fatty acid profiling, and gene expression analysis of key desaturases and elongases of the P. dumerilii LC-PUFA biosynthesis. Principal C omponent A nalysis (PCA) was conducted to explore patterns in fatty acid profiles and assess potential differences among treatments.
Results
For T rial 1, P. dumerilii exhibited elevated levels of ω3 LC-PUFA, specifically eicosatetraenoic acid (20:4n-3), eicosapentaenoic acid (20:5n-3), and docosapentaenoic acid (22:5n-3) at 40‰ salinity, compared to individuals maintained at 35‰ and 30‰, despite receiving the same diet. Gene expression analysis revealed a significant upregulation of t he P. dumerilii Δ6/Δ8 Fed desaturase under high salinity (40‰), accompanied by a decrease in its metabolic substrates (20:2n-6 and 20:3n-3); these results indicate an enhanced desaturase activity at high salinity . In Trial 2 , PCA of the fatty acid profiles showed a clear separation among the four experimental groups. The first component distinguished salinities, and the second separated diets with and without iron supplementation, especially at 40‰ salinity (Fig. 1). No effects from iron supplementation were observed at 35‰ salinity. Notably, at 40‰, iron supplementation correlated with increased levels of 20:3n-6 and 20:4n-3 (Fig. 1 ), both desaturase products from the P. dumerilii Δ6/Δ8 Fed and therefore supporting enhanced desaturase activity. These results support the conclusion that the ω3 LC-PUFA biosynthesis in P. dumerilii is influenced by both salinity and dietary iron. The proposed mechanism involves an upregulation of fatty acyl desaturases at high salinity, while iron supplementation guarantees the enzymatic activity that leads to enhanced LC-PUFA biosynthesis and improved nutritional value.
References
Monroig, Ó., Shu-Chien, A.C., Kabeya, N., Tocher, D.R., Castro, L.F.C., 2022. Desaturases and elongases involved in long-chain polyunsaturated fatty acid biosynthesis in aquatic animals: From genes to functions. Prog. Lipid Res. 86, 101157.
Palacios, E., Racotta, I.S., 2007. Salinity stress test and its relation to future performance and different physiological responses in shrimp post larvae. Aqua. 268, 123–135.
Shanklin J., Guy J.E., Mishra G., Lindqvist Y., 2009. Desaturases: Emerging m odels for u nderstanding f unctional d iversification of d iiron-containing enzymes. J. Biol. Chem. 84,18559–18563.
Acknowledgements
This research is part of the IMPROMEGA project RTI2018-095119-B-I00, funded by MCIN/ AEI /10.13039/501100011033/ and by FEDER, EU. Additionally, this study is part of the ThinkInAzul program, funded by the European Union NextGenerationEU (PRTR-C17. I1) and the Generalitat Valenciana (THINKINAZUL/2021/026). KB is funded by a predoctoral contract, Santiago Grisolía (GRISOLIA/2021/120), from the “Generalitat Valenciana”.