Creatine in giant grouper (Epinephelus lanceolatus): Insights into biosynthesis and transporter genes molecular characterization, regulation and impact on muscle metabolism

Abstract

Creatine is synthesized from glycine, arginine, and methionine with the catalysis of glycine amidinotransferase (GATM) and guanidinoacetate N-methyltransferase (GAMT), playing a crucial role in regulating the cellular metabolism and whole-body energy balance. However, information regarding the molecular characteristics of GATM and GAMT is limited in fish species. In this study, GATM and GAMT cDNA were cloned from giant grouper (Epinephelus lanceolatus). The cloned cDNA sequence of GATM was 1503 bp encoding a 421 aa containing conserved regions of amidinotransferase superfamily. GAMT cDNA was 892 bp encoding a 235 aa containing conserved regions of S-adenosylmethionine-dependent methyltransferases superfamily. The phylogenetic tree indicated that GATM and GAMT were clustered distinctly into teleost and featured common function domains and homology with teleost proteins, but they were distinct from the terrestrial clusters. The mRNA transcripts of GATM and GAMT were detectable mainly in muscle tissue. Additionally, as the creatine synthesis limited enzyme, GATM gene and protein expression were both significantly regulated by supplement of dietary creatine (P < 0.05). Metabolomics analysis showed that dietary creatine significantly affected skeletal muscle nutrients and energy metabolism. Meanwhile, inclusion of dietary creatine led to a significant inhibition of AMPK phosphorylation. What is more, we also recorded that cytochrome oxidase (cox) and mitochondrial fission factor (mff) gene expression were remarkable up-regulated by dietary creatine supplementation. In summary, the results obtained from the present study would help us better understand the creatine synthesis pathway in fish species. We also demonstrated that creatine plays a significant role in fish muscle metabolism.