Glutathione S-transferase pi (GSTpi) protects cells from death by altering intracellular oxidative stress. In order to understand the mechanism of GSTpi protection, a cell death model induced by serum depletion as the stress was established. Cotransfection of apoptosis signal-regulating kinase 1 (ASK1) and GSTpi cDNA was performed to elucidate the impact of GSTpi on ASK1 activity, as well as on its downstream signals, MKK7 and JNK, and to elucidate the potential protection of GSTpi on 293 cell death induced by serum depletion. The dominant negative mutant of JNK was used to explore if the blocking of the JNK pathway led to cell death inhibition. It was found that GSTpi had a dose-dependent inhibitory effect on activation induced by serum depletion, and also on inhibition both on MKK7 and JNK. Intracellular expression of GSTpi significantly inhibited serum depletion-induced cell death. Blocking the JNK pathway by transfection of a dominant negative form of JNK (JNK (APF)) brought about significant inhibition of cell death induced by serum depletion with an inhibiting rate as high as 15%. All the results suggest that the mechanism of GSTpi protection on serum depletion-induced cell death works through an ASK1-MKK7-JNK pathway.