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Structure–activity relations of leucine derivatives reveal critical moieties for cellular uptake and activation of mTORC1-mediated signaling

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Abstract

Among amino acids, leucine is a potential signaling molecule to regulate cell growth and metabolism by activating mechanistic target of rapamycin complex 1 (mTORC1). To reveal the critical structures of leucine molecule to activate mTORC1, we examined the structure–activity relationships of leucine derivatives in HeLa S3 cells for cellular uptake and for the induction of phosphorylation of p70 ribosomal S6 kinase 1 (p70S6K), a downstream effector of mTORC1. The activation of mTORC1 by leucine and its derivatives was the consequence of two successive events: the cellular uptake by l-type amino acid transporter 1 (LAT1) responsible for leucine uptake in HeLa S3 cells and the activation of mTORC1 following the transport. The structural requirement for the recognition by LAT1 was to have carbonyl oxygen, alkoxy oxygen of carboxyl group, amino group and hydrophobic side chain. In contrast, the requirement for mTORC1 activation was more rigorous. It additionally required fixed distance between carbonyl oxygen and alkoxy oxygen of carboxyl group, and amino group positioned at α-carbon. l-Configuration in chirality and appropriate length of side chain with a terminal isopropyl group were also important. This confirmed that LAT1 itself is not a leucine sensor. Some specialized leucine sensing mechanism with rigorous requirement for agonistic structures should exist inside the cells because leucine derivatives not transported by LAT1 did not activate mTORC1. Because LAT1–mTOR axis is involved in the regulation of cell growth and cancer progression, the results from this study may provide a new insight into therapeutics targeting both LAT1 and leucine sensor.

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Acknowledgments

The authors would like to thank Yasuhiro M. Umemura for his valuable advice and Yoko Tanaka for her technical assistance. This work was supported in part by Grant-in-Aid for Scientific Research and Grant-in-Aid for Young Scientists from Japan Society for the Promotion of Science (JSPS), by Regional Innovation Strategy Support Program, Grant-in-Aid for the Scientific Research on Innovative Areas “HD Physiology”, and Grant-in-Aid for Scientific Research on Priority Areas “Transportsome” from Ministry of Education, Culture, Sports, Science and Technology of Japan (MEXT), and by grants from the Osaka Medical Research Foundation for Intractable Diseases, the Advanced Research for Medical Products Mining Program of the National Institute of Biomedical Innovation, and the Ajinomoto Amino Acid Research Program.

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    1. Department of Bio-System Pharmacology, Graduate School of Medicine, Osaka University, 2-2 Yamadaoka, Suita, Osaka, 565-0871, Japan

      Shushi Nagamori, Pattama Wiriyasermkul, Suguru Okuda, Ryuichi Ohgaki & Yoshikatsu Kanai

    2. Department of Pharmaceutical Manufacturing Chemistry, Kyoto Pharmaceutical University, Kyoto, Japan

      Naoto Kojima

    3. Faculty of Pharmaceutical Sciences, Tokushima Bunri University, Tokushima, Japan

      Yoshiyuki Hari

    4. Department of Synthetic Chemistry and Biological Chemistry, Graduate School of Engineering, Kyoto University, Kyoto, Japan

      Shigeki Kiyonaka & Yasuo Mori

    5. Department of Technology and Ecology, Hall of Global Environmental Studies, Kyoto University, Kyoto, Japan

      Shigeki Kiyonaka & Yasuo Mori

    6. Advanced Clinical Research Center, Fukushima Medical University, Fukushima, Japan

      Hideyuki Tominaga

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    1. Shushi Nagamori
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    Correspondence to Yoshikatsu Kanai.

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    Handling Editor: E. I. Closs.

    S. Nagamori and P. Wiriyasermkul contributed equally to this work.

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    Nagamori, S., Wiriyasermkul, P., Okuda, S. et al. Structure–activity relations of leucine derivatives reveal critical moieties for cellular uptake and activation of mTORC1-mediated signaling. Amino Acids 48, 1045–1058 (2016). https://doi.org/10.1007/s00726-015-2158-z

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