Abstract
Glutathione peroxidase 1 (GPx-1) has been implicated in the etiology of several common diseases due to the association between specific allelic variations and cancer risk. The most common among these variations are the codon 198 polymorphism that results in either a leucine or proline and the number of alanine repeat codons in the coding sequence. The molecular and biologic consequences of these variations remain to be characterized. Toward achieving this goal, we have examined the cellular location of GPx-1 encoded by allelic variants by ectopically expressing these genes in MCF-7 human breast carcinoma cells that produce undetectable levels of GPx-1, thus achieving exclusive expression in the same cellular environment. A differential distribution between the cytoplasm and mitochondria was observed, with the allele expressing the leucine-198 polymorphism and 7 alanine repeats being more cytoplasmically located than the other alleles examined. To assess whether the distribution of GPx-1 between the cytoplasm and mitochondria had a biologic consequence, we engineered derivative GPx-1 proteins that were targeted to the mitochondria by the addition of a mitochondria targeting sequence and expressed these proteins in MCF-7 cells. These cells were examined for their response to oxidative stress, energy metabolism, and impact on cancer-associated signaling molecules. The results obtained indicated that both primary GPx-1 sequence and cellular location have a profound impact on cellular biology and offer feasible hypotheses about how expression of distinct GPx-1 alleles can affect cancer risk. Cancer Res; 74(18); 5118-26. ©2014 AACR.
Highlights
Glutathione peroxidase 1 (GPx-1) is the ubiquitously expressed and best characterized member of the GPx family of selenium-containing proteins that uses reducing equivalents from glutathione to detoxify peroxides
To assess the cellular location of the GPx-1 protein expressed by different alleles, cytoplasmic and mitochondrial cellular fractions were obtained from four previously generated MCF-7 transfectants containing either 5 or 7 alanine repeat codons (A5 or A7) and either a proline (P) or leucine (L) at codon 198, these being referred to as A5L, A7L, A5P, or A7P [9]
Consistent with several previous publications, GPx-1 was detected in both the cytoplasmic and mitochondrial fractions [20,21,22,23]. This analysis indicated that there was a differential distribution between the cytoplasm and the mitochondria among the proteins encoded by the distinct GPx-1 alleles, with A7L being less distributed to the mitochondria as compared with A5P, A5L, and A7P (Fig. 1A and B)
Summary
Glutathione peroxidase 1 (GPx-1) is the ubiquitously expressed and best characterized member of the GPx family of selenium-containing proteins that uses reducing equivalents from glutathione to detoxify peroxides (extensively reviewed in ref. 1). Glutathione peroxidase 1 (GPx-1) is the ubiquitously expressed and best characterized member of the GPx family of selenium-containing proteins that uses reducing equivalents from glutathione to detoxify peroxides The translation of GPx-1 includes the recognition of an in-frame UGA triplet as the codon for selenocysteine in a process that requires a selenocysteine insertion sequence in the 30-untranslated region of the GPx1 mRNA, a selenocysteine tRNA, and a host of selenoproteinspecific translation factors [2,3,4,5]. Support for a role for GPx-1 in cancer etiology comes from genetic data indicating significant associations between allelic variations in the GPx-1 gene and cancer risk [6, 7]. Two GPx-1 common genetic variations have been extensively studied; an.
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