PARENT SESSION
1:30 PM to 3:30 PM
Wednesday, April 24, 2002
Poster Session 35 Other
Room: Nevada Exhibition Center
(P39-372) Role of glutaredoxin in metabolic oxidative stress-activated ASK1-MEK-MAPK pathway.
Song, Jae*,1, Lee, Yong1, 1 University of Pittsburgh, Pittsburgh, PA
ABSTRACT-
We have previously observed that glucose deprivation increases the intracellular levels of hydroperoxide and oxidized glutathione. The increased steady state levels of hydroperoxide activate signal transduction pathways, such as activated protein kinase (SAPK). In this study, we further investigated how metabolic oxidative stress triggers signal transduction pathways in mammalian cells. We hypothesized that glutaredoxin (GRX), a redox-sensing molecule, interacts with glutathione disulfide and forms an intramolecular disulfide form of GRX (GRX-[S=S]). GRX-[S=S] dissociates from apoptosis signal-regulating kinase 1 (ASK1) leading to activation of the ASK1-MEK-MAPK signal transduction pathway. To examine whether GRX dissociates from ASK1 during glucose deprivation, multi drug-resistant human breast carcinoma MCF-7/ADR cells were co-infected adenoviral vectors containing His-tagged GRX and HA-tagged ASK1. Cells were exposed to glucose-free medium. Cell lysates were immunoprecipitated with anti-His antibody, and then immunoblotted with anti-HA antibody or anti-His antibody. Our data reveals that GRX binds to ASK1 in the presence of glucose. However, GRX is dissociated from ASK1 during glucose deprivation. These data suggest that the interaction between GRX and ASK1 is dependent upon the redox state of GRX. We further examine the interaction site of GRX with ASK1. Data with a series of deletion mutant assay suggest that GRX binds to the C-terminal portion of ASK1. Immune complex kinase assay reveals that MEK4 and JNK1 activity were promoted through ASK1 kinase activation during glucose deprivation. Our results demonstrate that GRX is a negative regulator of ASK1 and disrupting interaction between GRX and ASK1 activates the ASK1-MEK-MAPK signal transduction pathway.
KEYWORDS: glutaredoxin, oxidative stress, redox state, ASK1-MEK-MAPK