title: Eel green fluorescent protein is associated with resistance to oxidative stress.
abstract: Green fluorescent protein (GFP) from eel (Anguilla japonica) muscle (eelGFP) is unique in the vertebrates and requires bilirubin as a ligand to emit fluorescence. This study was performed to clarify the physiological function of the unique GFP. Investigation of susceptibility to oxidative stress was carried out using three types of cell lines including jellyfish (Aequorea coerulescens) GFP (jfGFP)-, or eel GFP (eelGFP)-expressing HEK293 cells, and control vector-transfected HEK293 cells. Binding of eelGFP to bilirubin was confirmed by the observation of green fluorescence in HEK293-eelGFP cells. The growth rate was compared with the three types of cells in the presence or absence of phenol red which possessed antioxidant activity. The growth rates of HEK293-CV and HEK293-jfGFP under phenol red-free conditions were reduced to 52 and 31% of those under phenol red. Under the phenol red-free condition, HEK293-eelGFP had a growth rate of approximately 70% of the phenol red-containing condition. The eelGFP-expressing cells were approximately 2-fold resistant to oxidative stress such as H2O2 exposure. The fluorescence intensity partially decreased or disappeared after exposure to H2O2, and heterogeneous intensity of fluorescence was also observed in isolated eel skeletal muscle cells. These results suggested eelGFP, but not jfGFP, coupled with bilirubin provided the antioxidant activity to the cells as compared to non-bound free bilirubin.
authors.affiliation: Laboratory of Marine Biotechnology, Faculty of Fisheries, Kagoshima University, 4-50-20 Shimoarata, Kagoshima 890-0056, Japan.
authors.forename: Aki
authors.initials: A
authors.lastname: Funahashi
authors.affiliation: Division of Food and Chemical Biology, Faculty of Fisheries, Kagoshima University, 4-50-20 Shimoarata, Kagoshima 890-0056, Japan. Electronic address: komatsu@fish.kagoshima-u.ac.jp.
authors.forename: Masaharu
authors.initials: M
authors.lastname: Komatsu
authors.affiliation: Department of Molecular Oncology, Graduate School of Medical and Dental Sciences, Kagoshima University, Kagoshima 890-8544, Japan; Center for the Research of Advanced Diagnosis and Therapy of Cancer, Graduate School of Medical and Dental Sciences, Kagoshima University, Kagoshima 890-8544, Japan.
authors.forename: Tatsuhiko
authors.initials: T
authors.lastname: Furukawa
authors.affiliation: Division of Food and Chemical Biology, Faculty of Fisheries, Kagoshima University, 4-50-20 Shimoarata, Kagoshima 890-0056, Japan.
authors.forename: Yuki
authors.initials: Y
authors.lastname: Yoshizono
authors.affiliation: Division of Food and Chemical Biology, Faculty of Fisheries, Kagoshima University, 4-50-20 Shimoarata, Kagoshima 890-0056, Japan.
authors.forename: Hikari
authors.initials: H
authors.lastname: Yoshizono
authors.affiliation: Division of Food and Chemical Biology, Faculty of Fisheries, Kagoshima University, 4-50-20 Shimoarata, Kagoshima 890-0056, Japan.
authors.forename: Yasuhiro
authors.initials: Y
authors.lastname: Orikawa
authors.affiliation: Division of Food and Chemical Biology, Faculty of Fisheries, Kagoshima University, 4-50-20 Shimoarata, Kagoshima 890-0056, Japan; Department of Domestic Science, Kagoshima Women's College, Kagoshima 890-8565, Japan.
authors.forename: Shota
authors.initials: S
authors.lastname: Takumi
authors.affiliation: Division of Food and Chemical Biology, Faculty of Fisheries, Kagoshima University, 4-50-20 Shimoarata, Kagoshima 890-0056, Japan.
authors.forename: Kazuhiro
authors.initials: K
authors.lastname: Shiozaki
authors.affiliation: Division of Food and Chemical Biology, Faculty of Fisheries, Kagoshima University, 4-50-20 Shimoarata, Kagoshima 890-0056, Japan.
authors.forename: Seiichi
authors.initials: S
authors.lastname: Hayashi
authors.affiliation: Laboratory of Marine Biotechnology, Faculty of Fisheries, Kagoshima University, 4-50-20 Shimoarata, Kagoshima 890-0056, Japan.
authors.forename: Yoshio
authors.initials: Y
authors.lastname: Kaminishi
authors.affiliation: Laboratory of Marine Biotechnology, Faculty of Fisheries, Kagoshima University, 4-50-20 Shimoarata, Kagoshima 890-0056, Japan.
authors.forename: Takao
authors.initials: T
authors.lastname: Itakura
mesh_terms: D000811:Anguilla / Q000502:physiology*; D000818:Animals; D001663:Bilirubin / Q000378:metabolism; D049109:Cell Proliferation / Q000235:genetics; D003001:Cloning, Molecular; D029941:Fish Proteins / Q000235:genetics* / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000235:genetics* / Q000378:metabolism; D057809:HEK293 Cells / Q000187:drug effects; D006801:Humans; D006861:Hydrogen Peroxide / Q000378:metabolism; D018384:Oxidative Stress
chemical_list: D029941:Fish Proteins; C485192:green fluorescent protein, Aequorea victoria; D049452:Green Fluorescent Proteins; D006861:Hydrogen Peroxide; D001663:Bilirubin
keywords: Anguilla japonica; Bilirubin; Eel; EelGFP; Oxidative stress