Safeguarding genome integrity: the checkpoint kinases ATR, CHK1 and WEE1 restrain CDK activity during normal DNA replication

Abstract

Mechanisms that preserve genome integrity are highly important during the normal life cycle of human cells. Loss of genome protective mechanisms can lead to the development of diseases such as cancer. Checkpoint kinases function in the cellular surveillance pathways that help cells to cope with DNA damage. Importantly, the checkpoint kinases ATR, CHK1 and WEE1 are not only activated in response to exogenous DNA damaging agents, but are active during normal S phase progression. Here, we review recent evidence that these checkpoint kinases are critical to avoid deleterious DNA breakage during DNA replication in normal, unperturbed cell cycle. Possible mechanisms how loss of these checkpoint kinases may cause DNA damage in S phase are discussed. We propose that the majority of DNA damage is induced as a consequence of deregulated CDK activity that forces unscheduled initiation of DNA replication. This could generate structures that are cleaved by DNA endonucleases leading to the formation of DNA double-strand breaks. Finally, we discuss how these S phase effects may impact on our understanding of cancer development following disruption of these checkpoint kinases, as well as on the potential of these kinases as targets for cancer treatment.

Figure 1.

Regulation of CDK activity by ATR, CHK1 and WEE1 determines replication initiation during normal S phase. CDK activity is negatively regulated by phosphorylation of the Tyrosine 15 residue. This residue is phosphorylated by WEE1 and dephosphorylated by CDC25. CDC25 is negatively regulated by CHK1, which in turn is stimulated by the ATR kinase.

Figure 2.

Model describing potential mechanisms how loss of checkpoint kinases may cause DNA breakage during normal S phase. Loss of WEE1, ATR or CHK1 leads to increased CDK activity, which causes unscheduled replication initiation and subsequent DNA breakage. Following massive unscheduled initiation of replication, aberrant fork structures accumulate that are prone to breakage resulting in DNA DSBs. The high CDK activity may activate nucleases that directly catalyse the formation of DNA DSBs.

Figure 3.

Model depicting how inhibition of checkpoint kinases may lead to double-strand breaks after replication fork reversal. When checkpoint kinases are inhibited CDK activity is up-regulated leading to increased origin firing. As a consequence of the enhanced origin activity, replication factors such as polymerase subunits may become limiting leading to fork stalling. Helicases can process stalled forks leading to fork reversal, which generates substrates for DNA endonucleases such as MUS81. Fork reversal may also occur due to increased initiation of replication that generates enhanced torsional stress due to helicase unwinding of the double-stranded DNA. This stress can be relieved by fork reversal, thereby forming substrates for endonuclease cleavage.

Figure 4.

S phase effects during cancer treatment with inhibitors of ATR, CHK1 or WEE1 kinases may promote cancer specific killing. Cancer cells often contain elevated CDK activity due to genetic alterations, which may induce replication failures leading to DNA damage in S phase and subsequent genomic instability and tumour progression. Treatment with inhibitors of ATR, CHK1 or WEE1 further increases CDK activity leading to massive CDK-mediated DNA damage in S phase and subsequent cell death of S phase cancer cells.