Received for publication June 28, 2007.
Revised July 20, 2007.
Accepted for publication July 23, 2007.

Regulation of glutathione synthesis via interaction between GTRAP3-18 and EAAC1 at plasma membrane

Abstract

Regulation of the cysteine transporter EAAC1 for intracellular glutathione (GSH) content was investigated using HEK293 cells as a model system. GSH content was significantly reduced by L-aspartate--hydroxamate (50-250 µM), an inhibitor of both EAAC1 and GLT1, both of which are transporters to take up cysteine, while dihydrokainate (1-100 µM), a specific inhibitor of GLT1, failed to do so. This indicates that EAAC1 is involved in GSH content in HEK293 cells. We examined the effect of GTRAP3-18, which is capable of interacting with EAAC1. The GSH content decreased when the GTRAP3-18 protein level at the plasma membrane was increased by methyl--cyclodextrin (250 µM), rendering the cells more vulnerable to oxidative stress. Intracellular GSH increased when the GTRAP3-18 protein level at the plasma membrane was decreased by antisense oligonucleotides, rendering the cells more resistant to oxidative stress. Furthermore, we found that the increase in GSH content produced by stimulating protein kinase C, a translocator and activator of EAAC1, was inhibited by an increase in cell surface GTRAP3-18 protein. These results show GTRAP3-18 to negatively and dominantly regulate cellular GSH content via interaction with EAAC1 at the plasma membrane.

Key words: Glutathione, Oxidative stress/antioxidants, Oxidative stress

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