The effect of menadione on glutathione S-transferase A1 (GSTA1): c-Jun N-terminal kinase (JNK) complex dissociation in human colonic adenocarcinoma Caco-2 cells

Glutathione S-transferases (GSTs) act as modulators of mitogen-activated protein kinase signal transduction pathways via a mechanism involving protein–protein interactions. We have demonstrated that GSTA1 forms complexes with JNK and modifies JNK activation during cellular stress, but the factors that influence complex association and dissociation are unknown. We hypothesized that menadione causes dissociation of GSTA1–JNK complexes, activates JNK, and the consequences of menadione exposure depend on GSTA1 expression. We demonstrate that menadione causes GSTA1–JNK dissociation and JNK activation in preconfluent Caco-2 cells, whereas postconfluent cells are resistant to this effect. Moreover, preconfluent cells are more sensitive than postconfluent cells to menadione-induced cytotoxicity. Activation of JNK is transient since removal of menadione causes GSTA1 to re-associate with JNK reducing cytotoxicity. Over-expression and knockdown of GSTA1 did not alter JNK activation by menadione or sensitivity to menadione-induced cytotoxicity. These results indicate that GSTA1–JNK complex integrity does not affect the ability of menadione to activate JNK. N-acetyl cysteine prevents GSH depletion and blocks menadione-induced complex dissociation, JNK activation and inhibits menadione-induced cytotoxicity. JNK activation and inhibits menadione-induced cytotoxicity. The data suggest that the mechanism of menadione-induced JNK activation involves the production of reactive oxygen species, likely superoxide anion, and intracellular GSH levels play an important role in preventing GSTA1–JNK complex dissociation, subsequent JNK activation and induction of cytotoxicity.

► GSTA1 forms complexes with JNK in non-stressed cells.
► Menadione activates JNK signalling and mediates GSTA1–JNK complex dissociation.
► GSTA1 and glutathione prevent JNK activation, and stabilize GSTA1–JNK complexes.
► Mechanism of menadione-mediated JNK activation involves reactive oxygen species.
► GSH plays an important role in preventing GSTA1–JNK complex dissociation.