Effect And Mechanism Of Different Dietary Selenium Levels On Ubiquitin-mediated Protein Degradation In The Muscle Of Rainbow Trout

Selenium（Se）is an essential micronutrient for fish.The appropriate level of diet Se is essential for fish to maintain normal physiological functions.And Sedeficiency and excess are not conducive to the normal growth and development of fish.The growth performance of fish is closely related to the efficiency of protein deposition in skeletal muscle,and which is regulated by protein synthesis and degradation.Ubiquitin proteasome pathway（UPP）,as the main pathway of intracellular protein selective degradation,participates in more than 80%of skeletal muscle protein degradation.Under normal growth conditions,maintaining a lower activity of the ubiquitin-proteasome system is of great significance for promoting muscle growth.Previous studies have shown that dietary Se can regulate fish growth by affecting ubiquitin-mediated muscle protein degradation.However,the effects and the regulation mechanisms of dietary Se deficiency and excess on ubiquitin proteasome pathway need to be further explored.In the present study,five groups of diets with different Se levels（0,0.5,1,4 and 16 mg/kg）were formulated using yeast selenium（Se-yeast）as Se source,respectively.After 10-weeks of feeding trial,the effects of different dietary Se levels on growth performance and oxidative status of rainbow trout（Oncorhynchus mykiss）were studied,and the groups that represent dietary Se deficiency,adequacy and excess were selected.On this basis,the effects and potential molecular mechanism of dietary Se on ubiquitin-mediated protein degradation in the muscle of rainbow trout were further explored.The results showed as follows:1.Compared with other groups,4 mg/kg dietary Se significantly increased the weight gain rate,specific growth rate and feed efficiency of rainbow trout（P<0.05）,and the muscle crude protein content was significantly increased in this group（P<0.05）.In addition,the supplementation of dietary Se significantly promoted Se deposition in the rainbow trout tissues,with the increase of Se concentration,the total Se contents in the muscle and liver were increased and showed a dose-effect relationship（P<0.05）.Histomorphological observations found that the dietary Se significantly increased the ratio of muscle fibers diameter compared with the basal diet（0 mg/kg）,whereas 16 mg/kg dietary Se significantly decreased the ratio of large sized muscle fibers than 4 mg/kg dietary Se（P<0.05）.Meanwhile,4 mg/kg Se effectively enhanced the body antioxidant capacity,significantly down-regulated the expression of heat shock protein Hsp70 in the muscle and liver（P<0.05）,and activated the Keap1/Nrf2 signaling pathway,significantly up-regulated the expressions of antioxidant enzymes related genes including SOD2,CAT,GPX1a,GPX1b1,GPX4a1and GPX4b in the muscle and liver（P<0.05）.As a result,fish fed the diet with 4 mg/kg Se showed higher glycine peroxidase activity and lower contents of malondialdehyde and nitric oxide in serum,liver and muscle than fish in the other dietary groups（P<0.05）.Based on the above experimental results,the groups representing dietary Se deficiency,adequacy and excess were selected,their added levels of Se were 0 mg/kg,4 mg/kg and 16 mg/kg,respectively.2.On the basis of the previous experiment,this experiment fed rainbow trout with the diets of Se deficiency（0 mg/kg）,adequate（4 mg/kg）and excessive（16 mg/kg）for10 weeks.The results showed that 4 mg/kg dietary Se significantly improved the growth performance of rainbow trout（P<0.05）.Compared with adequate dietary Se,Se deficiency and excess accelerated the muscle protein degradation rate,which significantly increased the contents of 3-methylhistidine and tyrosine in the muscle（P<0.05）.In addition,Se deficiency and excess significantly improved the abundance of ubiquitinated protein,and affected the activity of proteasome by up-regulating the transcriptional levels of proteasome-related genes including psma3,psmd1 and psmd6（P<0.05）.And the gene and protein expression levels muscle-specific E3 ligase Mu RF1 and Atrogin-1 in the muscle of rainbow trout were significantly increased when fish fed with Se-deficient and Se-excessive diets.We further detected the upstream signaling pathways of the ubiquitin proteasome and found that:Se deficiency and excess significantly reduced the phosphorylation levels of Akt Ser⁴⁷³ and Fox O3a,which is indicated by a significant decrease in the ratio of p-Akt Ser⁴⁷³/Akt and p-Fox O3a/Fox O3a（P<0.05）.Moreover,Se deficiency and excess activated the NF-κB signaling pathway and significantly up-regulated the expressions of pro-inflammatory factors including NF-κB,IκBα2,TNF-αand IL-8.The phosphorylation level of IκBαand the nuclear localization of NF-κB were also significantly increased in the Se-deficient and Se-excessive groups（P<0.05）.In summary,the main conclusions of this study include:（1）compared with other groups,4 mg/kg dietary Se effectively promoted the muscle growth of rainbow trout.And 4 mg/kg dietary Se also up-regulated the expressions of antioxidant enzyme-related genes by activating the Keap1/Nrf2 signaling pathway and increased the activities of antioxidant enzymes to maintain the body in low oxidative stress status;（2）compared to adequate dietary Se（4 mg/kg）,dietary Se deficiency（0 mg/kg）and excess（16 mg/kg）increased the abundance of ubiquitinated proteins in the muscle,and up-regulated the expressions of Mu RF1 and Atrogin-1 by regulating Akt/Fox O3a and NF-κB signaling pathways,which aggravated the ubiquitin-mediated muscle protein degradation of rainbow trout.