Diallyl disulfide inhibits p34cdc2 kinase activity through changes in complex formation and phosphorylation

Previous studies from our laboratory demonstrated that diallyl disulfide (DADS), an oil-soluble allyl sulfur compound found in processed garlic, markedly suppressed p34(cdc2) kinase activity and induced a G(2)/M phase arrest in cultured human colon tumor (HCT-15) cells. The present studies reveal that suppression of p34(cdc2) kinase activity by DADS does not result from direct interactions with the protein, but through changes in factors influencing the formation and conversion of the enzyme to its active form. Flow cytometric analyses showed that the increased proportion of cells in the G(2)/M phase following DADS treatment was accompanied by an increase in cyclin B-1 protein expression. A temporal and dose-dependent response in cyclin B-1 expression was observed in cells treated with DADS. Western blot analysis revealed that 50 mu M DADS did not influence the quantity of p34(cdc2) protein expressed, but did decrease the amount associated with cyclin B-1 by 26% (P < 0.05). Exposure of unsynchronized cells to 25 or 50 mu M DADS caused a trend towards increased p34(cdc2) hyperphosphorylation (17 and 22%, respectively). Exposure of synchronized cells to 100 mu M DADS increased p34(cdc2) hyperphosphorylation by 15% (P < 0.05). Consistent with its ability to slightly increase the quantity of hyperphosphorylated p34(cdc2), DADS, 25 or 50 mu M, decreased cdc25C protein expression by 23 and 46%, respectively (P < 0.05). The present studies suggest that the ability of DADS to inhibit p34(cdc2) kinase activation occurs because of decreased p34(cdc2)/cyclin B-1 complex formation and modest p34(cdc2) hyperphosphorylation.