title: A glutathione peroxidase 4 (GPx4) homologue from southern bluefin tuna is a secreted protein: first report of a secreted GPx4 isoform in vertebrates.
abstract: The antioxidant enzyme glutathione peroxidase 4 (GPx4) is capable of reducing complex lipid hydroperoxides in addition to hydrogen peroxide and organic hydroperoxides. Mammals express three GPx4 isoforms that are targeted to nucleoli, mitochondria or cytosol via variable amino termini. To better understand the role of this important antioxidant enzyme in marine finfish, we determined the subcellular localisation of a GPx4 homologue from southern bluefin tuna (Thunnus maccoyii; SBT). We created constructs for the expression of the selenocysteine-to-cysteine mutant of SBT GPx4 (GPx4C) tagged with enhanced green fluorescent protein (EGFP), including or lacking a putative amino-terminal signal peptide, and expressed the fusion proteins in a fish cell line. Fluorescence microscopy revealed that the full-length GPx4C-EGFP fusion protein localised to the trans-Golgi, suggesting that tuna GPx4 may be directed to the secretory pathway. Anti-GFP immunoblotting of cell lysates and proteins from culture media showed that the secretion of SBT GPx4 into the culture medium required an amino-terminal signal peptide. According to available sequence data, the SBT GPx4 isoform studied here is representative of other piscine GPx4 isoforms, suggesting that the secretion of at least one GPx4 isoform may be common amongst teleost fish.
authors.affiliation: School of Biological Sciences, Flinders University, Bedford Park, SA 5042, Australia. peter.bain@csiro.au
authors.forename: Peter A
authors.initials: PA
authors.lastname: Bain
authors.forename: Kathryn A
authors.initials: KA
authors.lastname: Schuller
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D017931:DNA Primers / Q000235:genetics; D005470:Fluorometry; D005979:Glutathione Peroxidase / Q000235:genetics* / Q000378:metabolism*; D049452:Green Fluorescent Proteins; D015151:Immunoblotting; D007211:Indoles; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D000080662:Phospholipid Hydroperoxide Glutathione Peroxidase; D017422:Sequence Analysis, DNA; D013045:Species Specificity; D014413:Tuna / Q000378:metabolism*; D021601:trans-Golgi Network / Q000378:metabolism*
chemical_list: D017931:DNA Primers; D007211:Indoles; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; C007293:DAPI; D000080662:Phospholipid Hydroperoxide Glutathione Peroxidase; D005979:Glutathione Peroxidase