Oxidative stress activates the c-Jun N-terminal kinase (JNK) pathway. However, 
the exact mechanisms by which reactive oxygen species (ROS) activate JNK are 
unclear. We found that the ability of hydrogen peroxide (H(2)O(2)) to induce JNK 
activation varied in different cell types. Pyrrolidine dithiocarbamate (PDTC), a 
presumed antioxidant, induced JNK activation on its own and enhanced JNK 
activation by H(2)O(2) in many cell types, including Jurkat, HEK293, and LNCaP 
and Tsu-Pr1 prostate cancer cells. The activation of JNK by PDTC, in the 
presence or absence of exogenous H(2)O(2), was dependent on its chelating 
ability to metal ions, most likely copper ions. Despite the strong 
JNK-activating ability, H(2)O(2) plus PDTC did not induce significant activation 
of the upstream kinases, SEK1/MKK4 and MKK7. However, the JNK inactivation rate 
was slower in cells treated with H(2)O(2) plus PDTC compared with the rate in 
cells treated with ultraviolet C (UV-C). Treatment of H(2)O(2) plus PDTC 
significantly decreased the expression levels of a JNK phosphatase, M3/6 (also 
named hVH-5), but not the levels of other phosphatases (PP2A and PP4). In 
contrast, UV-C irradiation did not cause the down-regulation of M3/6. These 
results suggest that JNK activation by H(2)O(2) plus PDTC resulted from the 
down-regulation of JNK phosphatases. Our data also reveal a necessity to 
carefully evaluate the pharmacological and biochemical properties of PDTC.