Differential regulation of protein synthesis in skeletal muscle and liver of neonatal pigs by leucine through an mTORC1-dependent pathway

Agus Suryawan,Teresa A Davis,Hanh V Nguyen,Rosemarie D Almonaci

doi:10.1186/2049-1891-3-3

Abstract

Neonatal growth is characterized by a high protein synthesis rate that is largely due to an enhanced sensitivity to the postprandial rise in insulin and amino acids, especially leucine. The mechanism of leucine's action in vivo is not well understood. In this study, we investigated the effect of leucine infusion on protein synthesis in skeletal muscle and liver of neonatal pigs. To evaluate the mode of action of leucine, we used rapamycin, an inhibitor of mammalian target of rapamycin (mTOR) complex-1 (mTORC1). Overnight-fasted 7-day-old piglets were treated with rapamycin for 1 hour and then infused with leucine (400 μmol·kg-1·h-1) for 1 hour. Leucine infusion increased the rate of protein synthesis, and ribosomal protein S6 kinase 1 (S6K1) and eukaryotic initiation factor (eIF) 4E-binding protein-1 (4E-BP1) phosphorylation in gastrocnemius and masseter muscles (P < 0.05), but not in the liver. The leucine-induced stimulation of protein synthesis and S6K1 and 4E-BP1 phosphorylation were completely blocked by rapamycin, suggesting that leucine action is by an mTORC1-dependent mechanism. Neither leucine nor rapamycin had any effect on the activation of the upstream mTORC1 regulators, AMP-activated protein kinase and protein kinase B, in skeletal muscle or liver. The activation of eIF2α and elongation factor 2 was not affected by leucine or rapamycin, indicating that these two pathways are not limiting steps of leucine-induced protein synthesis. These results suggest that leucine stimulates muscle protein synthesis in neonatal pigs by inducing the activation of mTORC1 and its downstream pathway leading to mRNA translation.

Highlights

One of the hallmarks of the neonatal period is rapid growth, which is due to a high rate of protein synthesis [1]
We wished to determine whether inhibition of mTOR complex 1 (mTORC1) activation by rapamycin could suppress leucine-induced stimulation of protein synthesis in neonatal pigs
We found that rapamycin administration completely blocked leucine-induced protein synthesis in both muscle types (P < 0.05; Figure 1)

Summary

Introduction

One of the hallmarks of the neonatal period is rapid growth, which is due to a high rate of protein synthesis [1]. We further demonstrated that the feeding-induced stimulation of protein synthesis in most tissues is independently regulated by insulin and amino acids [3]. We found that leucine alone can stimulate protein synthesis in neonatal pigs and this effect decreases with development [4,5]. The main function of mTORC1 is to regulate mRNA translation by directly phosphorylating two down-stream substrates, ribosomal protein S6 kinase 1 (S6K1) and eukaryotic initiation factor (eIF) 4E-binding protein-1 (4E-BP1). The second complex, mTORC2, has been postulated to regulate the activation of protein kinase B (PKB) [6]. The exact mechanisms by which nutrients/amino acids, especially leucine, activate mTORC1 have been partly elucidated using cell culture systems [9]. Eukaryotic initiation factor 2 and elongation factor 2 (eEF2) pathways have been shown to be affected by amino acids/leucine [12,13]

Methods

Results

Conclusion