7.7: DNA 损伤会使细胞周期停滞

为了应对 DNA 损伤，细胞可以暂停细胞周期以评估和修复断裂。 然而，细胞必须在细胞周期的某些关键阶段检查 DNA。 如果细胞周期在 DNA 复制之前暂停，细胞中的 DNA 含量将增加两倍。 另一方面，如果细胞在 DNA 复制之后、有丝分裂之前停滞，它们的 DNA 含量将是正常量的四倍。 由于有大量专门的蛋白可供使用，细胞必须在正确的时间使用正确的蛋白，以在严格调控的细胞周期中做出损伤反应。

由受损 DNA 引起的复制应激会启动一条精心设计的蛋白途径，通过适当的修复机制对特定类型的损伤做出反应。 例如，可导致 DNA 双链断裂的电离辐射会激活 ATM 蛋白，从而启动一系列分子相互作用，其中涉及非同源末端连接、同源修复和核苷酸切除修复途径等修复机制。 ATM 和 ATR 等激酶对在不同时间尺度上运行的两个不同过程中的复制块做出反应：(i) 相对较快的翻译后修饰，例如下游激酶的磷酸化，最终导致 CDK 激活所需的细胞周期磷酸酶 CDC25 受到抑制（ii） ）较慢的转录调控，其中研究最充分的是 p53 的作用。

p53 是一种转录因子，可以调节在细胞周期停滞、细胞凋亡或衰老中发挥关键作用的蛋白的表达。 在健康细胞中，p53 维持在较低浓度。 在检测到双链断裂后，ATM 通过磷酸化激活 p53。 这导致 CDK 抑制剂 p21 以及促凋亡 BAX 和 PUMA 蛋白的表达。 p21 通过抑制细胞周期蛋白-CDK 复合物来阻止细胞周期，该复合物使介导 G1 到 S 相转变的蛋白磷酸化。 因此，p53 对于 G1/S 检查点机制至关重要。 在p53突变或缺失的细胞中，细胞分裂不再受到调节，这种不受控制的细胞分裂会导致恶性肿瘤。 此外，p53 可以通过调节介导核苷酸切除修复和诱导 dNTP 合成的因子，直接激活修复途径，例如 NER。

At various checkpoints during the cell cycle, multiple enzymes probe the DNA for damage. To maintain the integrity of the genome, only intact, undamaged DNA is allowed to pass through this cycle, and on to the next generation. If DNA damage is detected, the cell cycle pauses until this is repaired.

During DNA replication in the S phase, helicase unwinds the DNA, and DNA polymerase synthesizes a new strand from the template - creating a Y-shaped structure called a replication fork. Damaged DNA stalls the replication fork, causing it to become unstable, and the helicase and polymerase to uncouple from the DNA.

To prevent the damaged single-stranded DNA from reannealing, replication protein A or RPA, coats the single-stranded DNA at the stalled replication fork. This complex is then detected by the ATR protein, also known as ataxia telangiectasia or Rad-3 related protein.

If the damaged DNA is not a single mutation but a full double strand break, a protein complex called MRN is recruited at the site, which bridges the two damaged ends of the DNA and provides a platform for binding of the ataxia-telangiectasia mutated, or ATM protein.

Both ATM and ATR are kinases, which means they catalyze the transfer of phosphate groups from phosphate-donating molecules such as NTPs, to specific substrates.

ATR and ATM phosphorylate the downstream kinases Chk1 and Chk2, respectively. Chk1 and Chk2 phosphorylate CDC25, which prevents it from accepting further phosphates from CDK1. CDK1 is the regulator of the cell cycle, and as long as it remains inactive, this prevents the cell from progressing to the S phase.

Another target of ATR and ATM phosphorylation is the transcription activator protein p53. Phosphorylated p53 can directly bind to DNA, which stimulates another gene to produce a protein called p21. p21 inhibits the cell division-stimulating protein cdk2, preventing the cell from progressing to the next stage of cell division.