Regulation of amino acid-sensitive TOR signaling by leucine analogues in adipocytes

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Abstract
In adipocytes, amino acids stimulate the target of rapamycin (TOR) signaling pathway leading to phosphorylation of the translational repressor, eIF‐4E binding protein‐I (4E‐BP1), and ribosomal protein S6. L‐leucine is the primary mediator of these effects. The structure‐activity relationships of a putative L‐leucine recognition site in adipocytes (LeuRA) that regulates TOR activity were analyzed by examining the effects of leucine analogues on the rapamycin‐sensitive phosphorylation of the translational repressor, eIF‐4E binding protein‐I (4E‐BP1), an index of TOR activity. Several amino acids that are structurally related to leucine strongly stimulated 4E‐BP1 phosphorylation at concentrations greater than the EC50 value for leucine. The order of potency was leucine > norleucine > threo‐L‐β‐hydroxyleucine ≃ Ile > Met ≃ Val. Other structural analogues of leucine, such as H‐α‐methyl‐D/L‐leucine, S‐(−)‐2‐amino‐4‐pentenoic acid, and 3‐amino‐4‐methylpentanoic acid, possessed only weak agonist activity. However, other leucine‐related compounds that are known agonists, antagonists, or ligands of other leucine binding/recognition sites did not affect 4E‐BP1 phosphorylation. We conclude from the data that small lipophilic modifications of the leucine R group and α‐hydrogen may be tolerated for agonist activity; however, leucine analogues with a modified amino group, a modified carboxylic group, charged R groups, or bulkier aliphatic R groups do not seem to possess significant agonist activity. Furthermore, the leucine recognition site that regulates TOR signaling in adipocytes appears to be different from the following: (1) a leucine receptor that regulates macroautophagy in liver, (2) a leucine recognition site that regulates TOR signaling in H4IIE hepatocytes, (3) leucyl tRNA or leucyl tRNA synthetase, (4) the gabapentin‐sensitive leucine transaminase, or (5) the system L‐amino acid transporter. J. Cell. Biochem. 77:234–251, 2000.