Branched-Chain Amino Acid Supplementation Alters the Abundance of Mechanistic Target of Rapamycin and Insulin Signaling Proteins in Subcutaneous Adipose Explants from Lactating Holstein Cows.

Yusheng Liang,Felipe C Cardoso,Claudia Parys,Fabiana F Cardoso,Juan J Loor

doi:10.3390/ani11092714

Abstract

Simple SummaryBranched-chain amino acids (BCAAs) are import regulators of mechanistic target of rapamycin (mTOR). In humans and rodents, increased circulating BCAA levels are positively associated with changes in protein abundance of insulin and amino acid (AA) signaling pathways in organs such as skeletal muscle and adipose. Unlike aspects of fatty acid metabolism (e.g., lipolysis, lipogenesis), it is unknown if BCAA directly affect subcutaneous adipose tissue (SAT) AA metabolism and insulin signaling. We propose that BCAA availability within SAT could enhance aspects of AA and insulin function by promoting increases in the abundance of key proteins.The objective of this study was to investigate changes in protein abundance of mTOR and insulin signaling pathway components along with amino acid (AA) transporters in bovine s.c. adipose (SAT) explants in response to increased supply of Leu, Ile, or Val. Explants of SAT from four lactating Holstein cows were incubated with high-glucose serum-free DMEM, to which the 10 essential AAs were added to create the following treatments: ideal mix of essential AA (IPAA; Lys:Met 2.9:1; Lys:Thr 1.8:1; Lys:His 2.38:1; Lys:Val 1.23:1; Lys:Ile 1.45:1; Lys:Leu 0.85:1; Lys:Arg 2.08:1) or IPAA supplemented with Ile, Val, or Leu to achieve a Lys:Ile of 1.29:1 (incIle), Lys:Val 1.12:1 (incVal), or Lys:Leu (incLeu) 0.78:1 for 4 h. Compared with IPAA, incLeu or incIle led to greater activation of protein kinase B (AKT; p-AKT/total AKT) and mTOR (p-mTOR/total mTOR). Total EAA in media averaged 7.8 ± 0.06 mmol/L across treatments. Incubation with incLeu, incIle, or incVal led to greater protein abundance of solute carrier family 38 member 1 (SLC38A1), a Gln transporter, and the BCAA catabolism enzyme branched-chain α-keto acid dehydrogenase kinase (BCKDK) compared with IPAA. Activation of eukaryotic elongation factor 2 (eEF2; p-eEF2/total eEF2) was also greater in response to incLeu, incIle, or incVal. Furthermore, compared with incLeu or incIle, incVal supplementation led to greater abundance of SLC38A1 and BCKDK. BCKDK is a rate-limiting enzyme regulating BCAA catabolism via inactivation and phosphorylation of the BCKD complex. Overall, data suggested that enhanced individual supplementation of BCAA activates mTOR and insulin signaling in SAT. Increased AA transport into tissue and lower BCAA catabolism could be part of the mechanism driving these responses. The potential practical applications for enhancing post-ruminal supply of BCAA via feeding in rumen-protected form support in vivo studies to ascertain the role of these AAs on adipose tissue biology.

Highlights

It is well-recognized in a number of mammalian species that mechanistic target of rapamycin, including two distinct protein complexes, mTOR complex 1and mTOR complex 2, regulates protein synthesis, cell growth, and proliferation [1,2,3]
MTOR complex 2, regulates protein synthesis, cell growth, and proliferation [1,2,3]. Both complexes are partly controlled by the supply of nutrients such as branched-chain amino acids (BCAA; Leu, Ile, and Val), underscoring the unique role that dietary compounds can have in essential cellular processes
Bovine adipose tissue has not been generally considered responsive to AA supply, recent research indicated that, compared with liver and skeletal muscle, the s.c. adipose tissue (SAT) in dairy cows has the greatest mRNA abundance of mitochondrial branched-chain aminotransferase (BCAT2) [8]; BCAA, Gln, and neutral AA transporters are expressed in bovine SAT [8,9]

Summary

Introduction

MTOR complex 2 (mTORC2), regulates protein synthesis, cell growth, and proliferation [1,2,3] Both complexes are partly controlled by the supply of nutrients such as branched-chain amino acids (BCAA; Leu, Ile, and Val), underscoring the unique role that dietary compounds can have in essential cellular processes. The recent demonstration via metabolomics analysis that a high body condition score is associated with greater BCAA degradation before calving [7] suggested there is a potential relationship between body fat and BCAA metabolism in dairy cows. Bovine adipose tissue has not been generally considered responsive to AA supply, recent research indicated that, compared with liver and skeletal muscle, the s.c. adipose tissue (SAT) in dairy cows has the greatest mRNA abundance of mitochondrial branched-chain aminotransferase (BCAT2) [8]; BCAA, Gln, and neutral AA transporters are expressed in bovine SAT [8,9]. Available data support the notion that bovine SAT is a potential site for BCAA uptake and metabolism

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