The ATMATRp53Mediated Pathway

The ATM(ATR)/p53 pathway plays a pivotal role in one of the checkpoint mechanisms that arrest the cell cycle at G1 phase following DNA damage (G1 checkpoint) (Fig. 3). As described in Subheading 3, ATM is activated in response to IR, whereas ATR is activated in response to replication inhibition or UV-induced damage. The activated ATM or ATR then phosphorylates p53 (on Ser-15), and this phosphorylation causes MDM2 to dissociate from p53, which stabilizes p53 and leads to its accumulation (128,129). Increased expression of ARF owing to E2F1 stabilization in response to DNA damage also blocks the inhibitory function of MDM2, thereby increasing the nuclear amount of p53.

The principal kinases relaying ATM(ATR)-initiated checkpoint signaling are preferentially CHK2 for ATM and CHK1 for ATR. In response to IR or DNA replication stress, ATR phosphorylates CHK1 at Ser-317 and Ser-345 (175,177,198), which moderately increases its kinase activity and allows it to propagate the signal to downstream effectors, including p53, which CHK1 phosphorylates on Ser-20 (129). In response to IR, ATM phosphorylates CHK2 at Thr-68 (199), followed by CHK2 autophosphorylation on Thr-383 and Thr-387 and the activation of several target proteins, including p53, which CHK2 also phosphorylates on Ser-20 (128,129). These Ser-20 phosphorylation events both induce MDM2 to dissociate from p53.

Spindle Checkpoint Pathway

Fig. 3. ATM(ATR)-mediated G1/S checkpoint pathways. DNA damage triggers a rapid cascade of phosphorylation events involving either the ATM and CHK2 (upon IR) or the ATR and CHK1 (upon UV light) kinases. These phosphorylation events activate the target protein kinases to trap and phosphorylate the next target proteins, thereby transmitting the DNA damage signals. It has been determined that in response to IR, ATM phosphorylates CHK2 at Thr-68, whereas ATR (or ATM) phosphorylates CHK1 at Ser-317 and Ser-345. In one pathway (left), the CHK2 or CHK1 kinase phosphorylates Cdc25A phosphatase at serines 75, 123, 178, 278, and 292 (100). Of these, the Ser-123 residue that is targeted by CHK2 and the Ser-75 residue that is phosphory-lated by CHK1 seem to be particularly critical residues of Cdc25A (202, 99). The phosphorylated Ser-123 or Ser-75 residue is recognized by the ubiquitination (Ub) enzyme, and this promotes the rapid degradation of Cdc25A by the proteasome. Due to the disappearance of Cdc25A phosphatase activity that this degradation causes, the CDK2/cyclin E complex is locked in its inactive form because of the presence of the inhibitory phosphorylation on the Thr-14 and Tyr-15 residues of CDK2. Thus, the CDK2/cyclin E complex fails to load Cdc45 onto chromatin and the blockade of the initiation of the DNA replication origins is maintained.

The stabilized and activated p53 protein that results from CHK1/CHK2-mediated phosphorylation induces the transcription of a large number of genes, including p21WAF1, which silences the kinase activities of the CDK2/cyclin E, CDK2/cyclin A, or CDK4(6)/cyclin D complexes. This prevents the complexes from loading the Cdc45 origin binding factor onto chromatin, which precludes the recruitment of DNA polymerases, thereby blocking initiation of DNA replication from the unfired origins (200,201). Another important consequence of inhibiting both the CDK2 and CDK4(6) kinase complexes is that these complexes cannot then phosphorylate pRB, which allows pRB to maintain its inhibition of the E2F-dependent transcription of S-phase genes that are essential for S-phase entry as described in Subheading 2.1.4. These effects all result in G1 arrest. Maintenance of the G1/S arrest by way of this pathway after DNA damage is a delayed response that requires the transcription, translation, and/or protein stabilization of key checkpoint transducers. However, once initiated, this pathway provides a long-lasting G1 arrest, and the entry into S phase is prevented as long as a single unrepaired DNA lesion is detected by the checkpoint machinery.

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  • oliver dickson
    What are chk1 chk2 kinases?
    8 years ago

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