Replication factory activation can be decoupled from the replication timing program by modulating Cdk levels

• Published in: The Journal of cell biology 2010-01, Vol.188 (2), p.209-221
• Main Authors: Thomson, Alexander M, Gillespie, Peter J, Blow, J. Julian
• Format: Article
• Language: English
• Subjects: Animals; Biochemistry; CDC2 Protein Kinase - genetics; CDC2 Protein Kinase - metabolism; Cell Cycle - genetics; CHO Cells; Chromosomes; Cricetinae; Cricetulus; Cyclins; Deoxyribonucleic acid; DNA; DNA - biosynthesis; DNA replication; DNA Replication - genetics; Eggs; Embryonic cells; Enzyme Activation - genetics; Genes, cdc - physiology; Incubation; Interphase; Kinases; Mitosis - genetics; Ova; Replication origin; Replicon; S Phase - genetics; Stress, Physiological - genetics; Time Factors; Xenopus laevis
• ISSN: 0021-9525; 1540-8140
• DOI: 10.1083/jcb.200911037

In the metazoan replication timing program, clusters of replication origins located in different subchromosomal domains fire at different times during S phase. We have used Xenopus laevis egg extracts to drive an accelerated replication timing program in mammalian nuclei. Although replicative stress caused checkpoint-induced slowing of the timing program, inhibition of checkpoint kinases in an unperturbed S phase did not accelerate it. Lowering cyclin-dependent kinase (Cdk) activity slowed both replication rate and progression through the timing program, whereas raising Cdk activity increased them. Surprisingly, modest alteration of Cdk activity changed the amount of DNA synthesized during different stages of the timing program. This was associated with a change in the number of active replication factories, whereas the distribution of origins within active factories remained relatively normal. The ability of Cdks to differentially effect replication initiation, factory activation, and progression through the timing program provides new insights into the way that chromosomal DNA replication is organized during S phase.