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Regulation of amino acid-sensitive TOR signaling by leucine analogues in adipocytes
1Department of Cellular and Molecular Physiology, Pennsylvania State University College of Medicine, Hershey, Pennsylvania 17033, USA. clynch@psu.edu
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 (LeuR(A)) 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 EC(50) value for leucine. The order of potency was leucine > norleucine > threo-L-beta-hydroxyleucine approximately Ile > Met approximately Val. Other structural analogues of leucine, such as H-alpha-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 alpha-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.
Insights
L-leucine activates the target of rapamycin (TOR) signaling pathway in adipocytes. Researchers identified key structural features of leucine necessary for activating TOR signaling, differentiating this site from other leucine-binding proteins.
Area of Science:
- Cellular signaling
- Molecular biology
- Biochemistry
Background:
- Amino acids, particularly L-leucine, stimulate the target of rapamycin (TOR) signaling pathway in adipocytes.
- TOR signaling regulates crucial cellular processes, including protein synthesis, via phosphorylation of targets like 4E-BP1 and S6.
- Understanding the specific L-leucine recognition site (LeuR(A)) is key to elucidating nutrient-sensing mechanisms in adipocytes.
Purpose of the Study:
- To analyze the structure-activity relationships of the L-leucine recognition site (LeuR(A)) in adipocytes that regulates TOR activity.
- To determine which structural modifications of L-leucine analogues affect their ability to stimulate 4E-BP1 phosphorylation, an indicator of TOR activity.
Main Methods:
- Examined the effects of various L-leucine analogues on the rapamycin-sensitive phosphorylation of 4E-BP1 in adipocytes.
- Assessed the potency of different amino acids and leucine analogues in stimulating 4E-BP1 phosphorylation.
Main Results:
- Several leucine-related amino acids, including norleucine and isoleucine, stimulated 4E-BP1 phosphorylation, with potency order: leucine > norleucine > threo-L-beta-hydroxyleucine ≈ Ile > Met ≈ Val.
- Weak agonist activity was observed for analogues with modifications to the alpha-hydrogen or small lipophilic R groups.
- Analogues with modified amino/carboxylic groups, charged R groups, or bulkier aliphatic R groups showed no significant agonist activity.
Conclusions:
- Small lipophilic modifications to leucine's R group and alpha-hydrogen are tolerated for agonist activity at LeuR(A).
- The LeuR(A) site appears distinct from other known leucine-binding sites, including those involved in macroautophagy, hepatic TOR signaling, and amino acid transport.