Analysis of Phospholipid Hydroperoxide Glutathione Peroxidase mRNA
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Phospholipid hydroperoxide glutathione peroxidase (PHGPX or GPX4, E.C. 1.11.1.12) is one of the four identified selenium-dependent glutathione peroxidases (GPX) in mammals (1 ). Both of the pig (2.8 kb) and the mouse (4.0 kb) GPX4 genes contain seven exons and six introns, with putative regulatory elements or binding sites for transcriptional factors (2 ,3 ). There is 95% homology among amino acid sequences deduced from the GPX4 cDNA of rat (4 ), mouse (5 ) and human (6 ). In contrast, the homology between GPX1 and GPX4 is less than 40%. There are two forms of GPX4: the long form (23 kDa) with a leader sequence for transportation to mitochondria, and the short form (20 kDa) or the non-mitochondrial form (7 ). Although GPX4 was initially considered mainly an antioxidant enzyme by reducing phospholipid hydroperoxides (8 ), it may be involved in sperm maturation (9 ) as there are abundant GPX4 activity and mRNA in testis (10 ,11 ) and rat epididymal spermatozoa (12 ). It exists as a soluble peroxidase in spermatids, but loses its activity in mature spermatozoa and persists as an oxidatively cross-linked insoluble protein (13 ). Besides, GPX4 is expressed in all major tissues studied so far. Comparatively, GPX4 mRNA and activity are less affected by changes in tissue selenium status than those of GPX1 or GPX3 (14 ). Because there is a selenium-independent enzyme that reduces phospholipid hydroperoxides (15 ), GPX4 mRNA analysis becomes a specific tool to distinguish these two enzymes for various biochemical and physiological studies.