In this study, NIH3T3 cells stably transfected with a cyclin B1-luciferase reporter vector were utilized to investigate if cyclin B1 promoter activity is linked to either DNA replication or the activities of various cyclin-cyclin dependent kinases (cdks). Synchronized cells treated at the time of serum re-stimulation with 2 micrograms/ml of the DNA synthesis inhibitor, aphidicolin, did not display an increase in luciferase activity in comparison to control cells. When treated with aphidicolin during S phase, luciferase activity decreased. In contrast, luciferase activity increased in cells treated at the time of serum re-stimulation with 200 microM olomoucine, a cyclin-cdk inhibitor. These results indicate that (1) cyclin B1 promoter activity in NIH3T3 cells is linked to a DNA replication checkpoint control mechanism; (2) the cyclin B1 gene can be activated in the absence of functional cyclin E-cdk2, cyclin A-cdk2, or cyclin B-cdk2; and (3) cyclin B1 gene activation can occur in G1 arrested cells under conditions in which the arrest is not directly linked to inhibition of DNA synthesis.
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