Staurosporine blocks cell progression through G1 between the cyclin D and cyclin E restriction points

The protein kinase inhibitor staurosporine (SSP) stops progression of normal nontransformed cells in the G1 phase of the cell cycle. This implies that at least one of the cell cycle associated kinases, essential for cell transit through G1, is sensitive to SSP. Using multivariate flow cytometry to correlate the expression of cyclin E or cyclin D with cellular DNA content (i.e., cell cycle position), we have presently characterized the point of action of SSP in relation to the expression of these cyclins. During stimulation of normal human lymphocytes by phytohemagglutinin, cyclin D was expressed early, peaking at 8-14 h, while cyclin E appeared later, reaching a maximum at the time of cell entrance to S phase (24 h). Addition of SSP at the time of cell stimulation, while markedly suppressing the expression of cyclin E, had a rather modest effect on the expression of cyclin D. The data indicate that the SSP sensitive kinase(s) involved in cell progression through G1 operate beyond the restriction point of cyclin D but prior to that of cyclin E. Thus, the target(s) of SSP is (are) either the p33cdk/cyclin E complex itself or other protein kinase(s), activated subsequent to the cyclin D but prior to the cyclin E restriction point, the activity of which is essential for cell transit through G1.