Lipid metabolism and m~6A RNA methylation are altered in lambs supplemented rumen-protected methionine and lysine in a low-protein diet

Background: Methionine or lysine has been reported to influence DNA methylation and fat metabolism, but their combined effects in N6-methyl-adenosine（m~6A） RNA methylation remain unclarified. The combined effects of rumen-protected methionine and lysine（RML） in a low-protein（LP） diet on lipid metabolism, m~6A RNA methylation,and fatty acid（FA） profiles in the liver and muscle of lambs were investigated. Sixty-three male lambs were divided into three treatment groups, three pens per group and seven lambs per pen. The lambs were fed a 14.5% crude protein（CP） diet（adequate protein NP）, 12.5% CP diet（LP）, and a LP diet plus RML（LP + RML） for 60 d.Results: The results showed that the addition of RML in a LP diet tended to lower the concentrations of plasma leptin（P = 0.07）, triglyceride（P = 0.05）, and non-esterified FA（P = 0.08）. Feeding a LP diet increased the enzyme activity or m RNA expression of lipogenic enzymes and decreased lipolytic enzymes compared with the NP diet.This effect was reversed by supplementation of RML with a LP diet. The inclusion of RML in a LP diet affected the polyunsaturated fatty acids（PUFA）, n-3 PUFA, and n-6 PUFA in the liver but not in the muscle, which might be linked with altered expression of FA desaturase-1（FADS1） and acetyl-Co A carboxylase（ACC）. A LP diet supplemented with RML increased（P ＜ 0.05） total m~6A levels in the liver and muscle and were accompanied by decreased expression of fat mass and obesity-associated protein（FTO） and alk B homologue 5（ALKBH5）. The m RNA expressions of methyltransferase-like 3（METTL3） and methyltransferase-like 14（METTL14） in the LP + RML diet group were lower than those in the other two groups. Supplementation of RML with a LP diet affected only liver YTH domain family（YTHDF2） proteins（P ＜ 0.05） and muscle YTHDF3（P = 0.09）, which can be explained by limited m~6Abinding proteins that were mediated in m RNA fate.Conclusions: Our findings showed that the inclusion of RML in a LP diet could alter fat deposition through modulations of lipogenesis and lipolysis in the liver and muscle. These changes in fat metabolism may be associated with the modification of m~6A RNA methylation.