Lovastatin arrests CHO cells between the origin decision point and the restriction point

Asynchronously growing Chinese hamster ovary (CI-IO) cells treated with the pro-drug, beta -lactone ring form of lovastatin were arrested in G(1)-phas e, Subsequent removal of lovastatin resulted in the synchronous entry of ce lls into S-phase regardless of the presence of mevalonic acid. Lovastatin-a rrested cells contained hypophosphorylated retinoblastoma protein (Rb) and required serum mitogens to enter S-phase after lovastatin removal, indicati ng that cell-cycle arrest is prior to the restriction paint (R-point). Howe ver, in contrast to quiescent cells, intact nuclei prepared from lovastatin -arrested cells were competent for DNA replication when introduced into Xen opus egg extracts. Initiation of replication by Xenopus egg cytosol took pl ace specifically within the dihydrofolate reductase (DHFR) origin locus, de monstrating that cells were arrested after the origin decision point (ODP), We conclude that the beta -lactone ring form of lovastatin is an effective reagent with which to synchronize CHO cells between the ODP and R-point, w ithout resulting in the withdrawal of cells from the cell-cycle into a quie scent state. (C) 2000 Federation of European Biochemical Societies. Publish ed by Elsevier Science B.V. All rights reserved.