Establishment of Stable Cell Lines With High Expression of Heterodimers of Human 4F2hc and Human Amino Acid Transporter LAT1 or LAT2 and Delineation of Their Differential Interaction With α-Alkyl Moieties

Narakorn Khunweeraphong; Shushi Nagamori; Pattama Wiriyasermkul; Yumiko Nishinaka; Printip Wongthai; Ryuichi Ohgaki; Hidekazu Tanaka; Hideyuki Tominaga; Hiroyuki Sakurai; Yoshikatsu Kanai

doi:10.1254/jphs.12124FP

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Establishment of Stable Cell Lines With High Expression of Heterodimers of Human 4F2hc and Human Amino Acid Transporter LAT1 or LAT2 and Delineation of Their Differential Interaction With α-Alkyl Moieties

Narakorn Khunweeraphong, Shushi Nagamori, Pattama Wiriyasermkul, Yumiko Nishinaka, Printip Wongthai, Ryuichi Ohgaki, Hidekazu Tanaka, Hideyuki Tominaga, Hiroyuki Sakurai, Yoshikatsu Kanai

Author information

Narakorn Khunweeraphong
Division of Bio-system Pharmacology, Department of Pharmacology, Graduate School of Medicine, Osaka University, Japan
Department of Pharmacology and Toxicology, Kyorin University School of Medicine, Japan
Shushi Nagamori
Division of Bio-system Pharmacology, Department of Pharmacology, Graduate School of Medicine, Osaka University, Japan
Pattama Wiriyasermkul
Division of Bio-system Pharmacology, Department of Pharmacology, Graduate School of Medicine, Osaka University, Japan
Yumiko Nishinaka
Division of Bio-system Pharmacology, Department of Pharmacology, Graduate School of Medicine, Osaka University, Japan
Printip Wongthai
Division of Bio-system Pharmacology, Department of Pharmacology, Graduate School of Medicine, Osaka University, Japan
Ryuichi Ohgaki
Division of Bio-system Pharmacology, Department of Pharmacology, Graduate School of Medicine, Osaka University, Japan
Hidekazu Tanaka
Division of Bio-system Pharmacology, Department of Pharmacology, Graduate School of Medicine, Osaka University, Japan
Hideyuki Tominaga
Department of Molecular Imaging, Gunma University Graduate School of Medicine, Japan
Hiroyuki Sakurai
Department of Pharmacology and Toxicology, Kyorin University School of Medicine, Japan
Yoshikatsu Kanai
Division of Bio-system Pharmacology, Department of Pharmacology, Graduate School of Medicine, Osaka University, Japan

Keywords: transporter, amino acid, system L, blood–brain barrier, cancer

JOURNAL FREE ACCESS

2012 Volume 119 Issue 4 Pages 368-380

DOI https://doi.org/10.1254/jphs.12124FP

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Abstract

System L is a major transport system for cellular uptake of neutral amino acids. Among system L transporters, L-type amino acid transporter 1 (LAT1) is responsible for the nutrient uptake in cancer cells, whereas L-type amino acid transporter 2 (LAT2) is a transporter for non-cancer cells. In this study, we have established HEK293 cell lines stably expressing high levels of human LAT1 and LAT2 forming heterodimers with native human 4F2hc of the cells. We have found that L-[¹⁴C]alanine is an appropriate substrate to examine the function of LAT2, whereas L-[¹⁴C]leucine is used for LAT1. By using L-[¹⁴C]alanine on LAT2, we have for the first time directly evaluated the function of human LAT2 expressed in mammalian cells and obtained its reliable kinetics. Using α-alkyl amino acids including α-methyl-alanine and α-ethyl-L-alanine, we have demonstrated that α-alkyl groups interfere with the interaction with LAT2. These cell lines with higher practical advantages would be useful for screening and analyzing compounds to develop LAT1-specific drugs that can be used for cancer diagnosis and therapeutics. The strategy that we took to establish the cell lines would also be applicable to the other heterodimeric transporters with important therapeutic implications.

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