Cleavage of GSK-3ß by calpain counteracts the inhibitory effect of Ser9 phosphorylation on GSK-3ß activity induced by H2O2.

ABSTRACT

Glycogen synthase kinase-3 beta (GSK-3ß) dysfunction may play an essential role in the pathogenesis of psychiatric, metabolic, neurodegenerative diseases, in which oxidative stress exists concurrently. Some studies have shown that GSK-3ß activity is up-regulated under oxidative stress. This study evaluated how oxidative stress regulates GSK-3ß activity in human embryonic kidney 293 (HEK293)/Tau cells treated with hydrogen peroxide (H2O2). Here, we show that H2O2 induced an obvious increase of GSK-3ß activity. Surprisingly, H2O2 dramatically increased phosphorylation of GSK-3ß at Ser9, an inactive form of GSK-3ß,while there were no changes of phosphorylation of GSK-3ß at Tyr216. Moreover, H2O2 led to a transient [Ca²âº](i) elevation, and simultaneously increased the truncation of GSK-3ß into two fragments of 40 kDa and 30 kDa, whereas inhibition of calpain decreased the truncation and recovered the activity of GSK-3ß. Furthermore, tau was hyperphosphorylated at Ser396, Ser404, and Thr231, three most common GSK-3ß targeted sites after 100 µM H2O2 administration in HEK293/Tau cells, whereas inhibition of calpain blocked the tau phosphorylation. In addition, we found that there were no obvious changes of Cyclin-dependent kinase 5 (CDK5) expression (responsible for tau phosphorylation) and of p35 cleavage, the regulatory subunit of CDK5 in H2O2-treated HEK293/Tau cells. In conclusion, Ca²âº-dependent calpain activation leads to GSK-3ß truncation, which counteracts the inhibitory effect of Ser9 phosphorylation, up-regulates GSK-3ß activity, and phosphorylates tau in H2O2-treated HEK293/Tau cells.