Potential role of leucine metabolism in the leucine-signaling pathway involving mTOR

Abstract

Leucine has been shown to stimulate adipose tissue protein synthesis in vivo as well as leptin secretion, protein synthesis, hyper-plastic growth, and tissue morphogenesis in in vitro experiments using freshly isolated adipocytes. Recently, others have proposed that leucine oxidation in the mitochondria may be required to activate the mammalian target of rapamycin (mTOR), the cytosolic Ser/Thr protein kinase that appears to mediate some of these effects. The first irreversible and rate-limiting step in leucine oxidation is catalyzed by the branched-chain α-keto acid dehydrogenase (BCKD) complex. The activity of this complex is regulated acutely by phosphorylation of the E1α-subunit at Ser293 (S293), which inactivates the complex. Because the α-keto acid of leucine regulates the activity of BCKD kinase, it has been suggested as a potential target for leucine regulation of mTOR. To study the regulation of BCKD phosphorylation and its potential link to mTOR activation, a phosphopeptide-specific antibody recognizing this site was developed and characterized. Phospho-S293 (pS293) immunoreactivity in liver corresponded closely to diet-induced changes in BCKD activity state. Immunoreactivity was also increased in TREMK-4 cells after the induction of BCKD kinase by a drug-inducible promoter. BCKD S293 phosphorylations in adipose tissue and gastrocnemius (which is mostly inactive in vivo) were similar. This suggests that BCKD complex in epididymal adipose tissue from food-deprived rats is mostly inactive (unable to oxidize leucine), as is the case in muscle. To begin to test the leucine oxidation hypothesis of mTOR activation, the dose-dependent effects of orally administered leucine on acute activation of S6K1 (an mTOR substrate) and BCKD were compared using the pS293 antibodies. Increasing doses of leucine directly correlated with increases in plasma leucine concentration. Phosphorylation of S6K1 (Thr389, the phosphorylation site leading to activation) in adipose tissue was maximal at a dose of leucine that increased plasma leucine approximately threefold. Changes in BCKD phosphorylation state required higher plasma leucine concentrations. The results seem more consistent with a role for BCKD and BCKD kinase in the activation of leucine metabolism/oxidation than in the activation of the leucine signal to mTOR.

Conversion of leucine to β-hydroxy-β-methylbutyrate by α-keto isocaproate dioxygenase is required for a potent stimulation of protein synthesis in L6 rat myotubes

“…Leucine is transaminated in muscle by branched‐chain aminotransferase to α‐ketoisocaproate. Then, α‐ketoisocaproate is decarboxylated to isovaleryl‐CoA by the mitochondrial enzyme branched‐chain α‐ketoacid dehydrogenase and finally acyl‐CoA derivatives are formed and entered into the citric acid cycle 36. The anticatabolic actions of L‐Leu metabolites such as α‐ketoisocaproate have been known for 35 years 47…”

Dietary melatonin supplementation alters uteroplacental amino acid flux during intrauterine growth restriction in ewes

“…BCAA have been shown to regulate mammalian target of rapamycin (mTOR), which has been implicated as a nutritional sensor, that regulates cell growth and protein synthesis via rates of gene transcription and mRNA translation (Lynch et al, 2003;Wullschleger et al, 2006). Moreover, supplementation of BCAA to dams on a protein deficient diet can restore fetal growth and minimize the decreases in fetal organ mass and carcass fat, which is associated with increased mTOR signaling (Teodoro et al, 2012).…”

Cytosolic Branched Chain Aminotransferase (BCATc) Regulates mTORC1 Signaling and Glycolytic Metabolism in CD4+ T Cells

“…BCAT Enzyme Activity-Total branched chain aminotransferase activity was measured in solubilized cells at 37°C in 25 mM potassium phosphate buffer, pH 7.8, using 1 mM ␣-keto [1][2][3][4][5][6][7][8][9][10][11][12][13][14] C]isovalerate and 12 mM Ile as described (7). A unit of enzyme activity was defined as 1 nmol of Val formed per min/mg of protein at 37°C.…”

“…Tight control of BCAA metabolism is also important because the BCAA Leu is a nutrient signal that promotes insulin release from the pancreas (11) and stimulates protein synthesis in skeletal muscle, heart, adipose tissue, and liver (12)(13)(14). The stimulatory effects of Leu on protein translation are mediated by mTOR (12,15,16).…”