Regulation of reactive oxygen species and stress fiber formation by calpeptin in Swiss 3T3 fibroblasts.
We have investigated a novel function of calpeptin, a commonly used inhibitor of calpain, in the production of intracellular reactive oxygen species (ROS) in Swiss 3T3 fibroblasts. Calpeptin induced a rapid increase of intracellular ROS by a dose-dependent manner, with a maximal increase at 10 min, which was inhibited by ROS scavengers, catalase and 2-MPG. However, other calpain inhibitors, E64d and N-acetyl-Leu-Leu-Nle-CHO (ALLN), had no effect on the level of intracellular ROS, indicating that calpain was not involved in the ROS production by calpeptin. The role of Rho in the ROS production by calpain was studied by scrape-loading of C3 transferase. C3 transferase, which inhibited stress fiber formation by calpeptin, had no effect on the ROS production in response to calpeptin, suggesting that Rho was not involved in the ROS production by calpeptin. But the elevation of intracellular ROS was inhibited by mepacrine, a phospholipase A2 inhibitor. In addition, scavenging intracellular ROS by the incubation with catalase and 2-MPG had no effect on the stress fiber formation by calpeptin. These results suggested that calpeptin stimulated the production of intracellular ROS and stress fiber formation by independent mechanisms.
Kim
Soo-Jung
SJ
Biomolecule Research Team, Korea Basic Science Institute, 305-333, Taejon, South Korea.
Lee
Zee-Won
ZW
Kweon
Soo-Mi
SM
Kim
Soohyun
S
Ha
Kwon-Soo
KS
D016475:3T3 Cells; D000818:Animals; D002154:Calpain / Q000037:antagonists & inhibitors*; D015853:Cysteine Proteinase Inhibitors / Q000378:metabolism* / Q000494:pharmacology; D004151:Dipeptides / Q000378:metabolism* / Q000494:pharmacology; D007424:Intracellular Fluid / Q000378:metabolism; D051379:Mice; D010741:Phospholipases A / Q000378:metabolism; D054467:Phospholipases A2; D017382:Reactive Oxygen Species / Q000378:metabolism*; D020741:rho GTP-Binding Proteins / Q000378:metabolism
D015853:Cysteine Proteinase Inhibitors; D004151:Dipeptides; D017382:Reactive Oxygen Species; C071482:calpeptin; D010741:Phospholipases A; D054467:Phospholipases A2; D002154:Calpain; D020741:rho GTP-Binding Proteins