Hydrogen peroxide induces association between glyceraldehyde 3-phosphate dehydrogenase and phospholipase D2 to facilitate phospholipase D2 activation in PC12 cells.

Abstract Oxidative stress or signaling is widely implicated in apoptosis, ischemia and mitogenesis. Previously, our group reported that the hydrogen peroxide (H[sub 2]O[sub 2])-dependent activation of phospholipase D2 (PLD2) in PC12 cells is involved in anti-apoptotic effect . However, the precise mechanism of PLD2 activation by H[sub 2]O[sub 2] was not revealed. To find H[sub 2]O[sub 2]-dependent PLD2-regulating proteins, we immunoprecipitated PLD2 from PC12 cells and found that glyceraldehyde 3-phosphate dehydrogenase (GAPDH) coimmunoprecipitated with PLD2 upon H[sub 2]O[sub 2] treatment. This interaction was found to be direct by in vitro reconstitution of purified GAPDH and PLD2. In vitro studies also indicated that PLD2-associated GAPDH was modified on its reactive cysteine residues. Koningic acid, an alkylator of GAPDH on catalytic cysteine residue, also increased interaction between the two proteins in vitro and enhanced PLD2 activity in PC12 cells. Blocking H[sub 2]O[sub 2]-dependent modification of GAPDH with 3-aminobenzamide resulted in the inhibition of the GAPDH/PLD2 interaction and attenuated H[sub 2]O[sub 2]-induced PLD2 activation in PC12 cells. From the results, we suggest that H[sub 2]O[sub 2] modifies GAPDH on its catalytic cysteine residue not only to inactivate the dehydrogenase activity of GAPDH but also to endow GAPDH with the ability to bind PLD2 and the resulting association is involved in the regulation of PLD2 activity by H[sub 2]O[sub 2]. [ABSTRACT FROM AUTHOR]