Calmodulin as a Potential Target by Which Berberine Induces Cell Cycle Arrest in Human Hepatoma Bel7402 Cells

Berberine is an isoquinoline alkaloid that has drawn extensive attention because it possesses various biological activities. Several mechanisms have been proposed to interpret the anticancer activity of berberine. However, these explanations are mostly based on its downstream-regulated genes or proteins; information on the direct target proteins that mediate the antiproliferative action of berberine remains unclear. In this study, a computational pipeline based on a ligandprotein inverse docking program and mining of the Connectivity MAP' data was adopted to explore the potential target proteins for berberine. The results showed that four proteins, that is calmodulin, cytochrome P450 3A4, sex hormone-binding globulin, and carbonic anhydrase II, were suggested to be the potential targets of berberine. The anticalmodulin property of berberine was demonstrated with an in vitro phosphodiesterase activity assay. Flow cytometric analysis found that G1 cell cycle arrest induced by berberine in Bel7402 cells was enhanced by cotreatment with calmodulin inhibitors. Western blotting results indicated that berberine treatment decreased phosphorylation of calmodulin kinase II and blocked subsequent MEK1 activation as well as p27 protein degradation. These results suggested that calmodulin might play crucial roles in berberine-induced cell cycle arrest in cancer cells.