NRFL-1, the C. elegans NHERF orthologue, interacts with amino acid transporter 6 (AAT-6) for age-dependent maintenance of AAT-6 on the membrane

Kohei Hagiwara, Shushi Nagamori, Yasuhiro M. Umemura, Ryuichi Ohgaki, Hidekazu Tanaka, Daisuke Murata, Saya Nakagomi, Kazuko H. Nomura, Eriko Kage-Nakadai, Shohei Mitani, Kazuya Nomura, Yoshikatsu Kanai

Research output: Contribution to journalArticle

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Abstract

The NHERF (Na+/H+ exchanger regulatory factor) family has been proposed to play a key role in regulating transmembrane protein localization and retention at the plasma membrane. Due to the high homology between the family members, potential functional compensations have been a concern in sorting out the function of individual NHERF numbers. Here, we studied C. elegans NRFL-1 (C01F6.6) (nherf-like protein 1), the sole C. elegans orthologue of the NHERF family, which makes worm a model with low genetic redundancy of NHERF homologues. Integrating bioinformatic knowledge of C. elegans proteins into yeast two-hybrid scheme, we identified NRFL-1 as an interactor of AAT-6, a member of the C. elegans AAT (amino acid transporter) family. A combination of GST pull-down assay, localization study, and co-immunoprecipitation confirmed the binding and characterized the PDZ interaction. AAT-6 localizes to the luminal membrane even in the absence of NRFL-1 when the worm is up to four-day old. A fluorescence recovery after photobleaching (FRAP) analysis suggested that NRFL-1 immobilizes AAT-6 at the luminal membrane. When the nrfl-1 deficient worm is six-day or older, in contrast, the membranous localization of AAT-6 is not observed, whereas AAT-6 tightly localizes to the membrane in worms with NRFL-1. Sorting out the in vivo functions of the C. elegans NHERF protein, we found that NRFL-1, a PDZ-interactor of AAT-6, is responsible for the immobilization and the age-dependent maintenance of AAT-6 on the intestinal luminal membrane.

Original languageEnglish
Article numbere43050
JournalPloS one
Volume7
Issue number8
DOIs
Publication statusPublished - Aug 15 2012

Fingerprint

amino acid transporters
Amino Acid Transport Systems
Maintenance
Membranes
Sorting
sorting
Caenorhabditis elegans Proteins
Fluorescence Recovery After Photobleaching
Photobleaching
transmembrane proteins
proteins
sodium-hydrogen exchanger regulatory factor
sodium-hydrogen antiporter
Cell membranes
Bioinformatics
Computational Biology
Immunoprecipitation
bioinformatics
Immobilization
Yeast

All Science Journal Classification (ASJC) codes

  • Biochemistry, Genetics and Molecular Biology(all)
  • Agricultural and Biological Sciences(all)

Cite this

NRFL-1, the C. elegans NHERF orthologue, interacts with amino acid transporter 6 (AAT-6) for age-dependent maintenance of AAT-6 on the membrane. / Hagiwara, Kohei; Nagamori, Shushi; Umemura, Yasuhiro M.; Ohgaki, Ryuichi; Tanaka, Hidekazu; Murata, Daisuke; Nakagomi, Saya; Nomura, Kazuko H.; Kage-Nakadai, Eriko; Mitani, Shohei; Nomura, Kazuya; Kanai, Yoshikatsu.

In: PloS one, Vol. 7, No. 8, e43050, 15.08.2012.

Research output: Contribution to journalArticle

Hagiwara, K, Nagamori, S, Umemura, YM, Ohgaki, R, Tanaka, H, Murata, D, Nakagomi, S, Nomura, KH, Kage-Nakadai, E, Mitani, S, Nomura, K & Kanai, Y 2012, 'NRFL-1, the C. elegans NHERF orthologue, interacts with amino acid transporter 6 (AAT-6) for age-dependent maintenance of AAT-6 on the membrane', PloS one, vol. 7, no. 8, e43050. https://doi.org/10.1371/journal.pone.0043050
Hagiwara, Kohei ; Nagamori, Shushi ; Umemura, Yasuhiro M. ; Ohgaki, Ryuichi ; Tanaka, Hidekazu ; Murata, Daisuke ; Nakagomi, Saya ; Nomura, Kazuko H. ; Kage-Nakadai, Eriko ; Mitani, Shohei ; Nomura, Kazuya ; Kanai, Yoshikatsu. / NRFL-1, the C. elegans NHERF orthologue, interacts with amino acid transporter 6 (AAT-6) for age-dependent maintenance of AAT-6 on the membrane. In: PloS one. 2012 ; Vol. 7, No. 8.
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abstract = "The NHERF (Na+/H+ exchanger regulatory factor) family has been proposed to play a key role in regulating transmembrane protein localization and retention at the plasma membrane. Due to the high homology between the family members, potential functional compensations have been a concern in sorting out the function of individual NHERF numbers. Here, we studied C. elegans NRFL-1 (C01F6.6) (nherf-like protein 1), the sole C. elegans orthologue of the NHERF family, which makes worm a model with low genetic redundancy of NHERF homologues. Integrating bioinformatic knowledge of C. elegans proteins into yeast two-hybrid scheme, we identified NRFL-1 as an interactor of AAT-6, a member of the C. elegans AAT (amino acid transporter) family. A combination of GST pull-down assay, localization study, and co-immunoprecipitation confirmed the binding and characterized the PDZ interaction. AAT-6 localizes to the luminal membrane even in the absence of NRFL-1 when the worm is up to four-day old. A fluorescence recovery after photobleaching (FRAP) analysis suggested that NRFL-1 immobilizes AAT-6 at the luminal membrane. When the nrfl-1 deficient worm is six-day or older, in contrast, the membranous localization of AAT-6 is not observed, whereas AAT-6 tightly localizes to the membrane in worms with NRFL-1. Sorting out the in vivo functions of the C. elegans NHERF protein, we found that NRFL-1, a PDZ-interactor of AAT-6, is responsible for the immobilization and the age-dependent maintenance of AAT-6 on the intestinal luminal membrane.",
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AU - Nagamori, Shushi

AU - Umemura, Yasuhiro M.

AU - Ohgaki, Ryuichi

AU - Tanaka, Hidekazu

AU - Murata, Daisuke

AU - Nakagomi, Saya

AU - Nomura, Kazuko H.

AU - Kage-Nakadai, Eriko

AU - Mitani, Shohei

AU - Nomura, Kazuya

AU - Kanai, Yoshikatsu

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