Unidad de Investigación, Hospital Universitario de Canarias, Instituto de Tecnologias Biomedicas, 38320 Tenerife, Spain.
J Cell Biol. 2011 Aug 22;194(4):567-79. doi: 10.1083/jcb.201101047.
Correct replication of the genome and protection of its integrity are essential for cell survival. In a high-throughput screen studying H2AX phosphorylation, we identified Wee1 as a regulator of genomic stability. Wee1 down-regulation not only induced H2AX phosphorylation but also triggered a general deoxyribonucleic acid (DNA) damage response (DDR) and caused a block in DNA replication, resulting in accumulation of cells in S phase. Wee1-deficient cells showed a decrease in replication fork speed, demonstrating the involvement of Wee1 in DNA replication. Inhibiting Wee1 in cells treated with short treatment of hydroxyurea enhanced the DDR, which suggests that Wee1 specifically protects the stability of stalled replication forks. Notably, the DDR induced by depletion of Wee1 critically depends on the Mus81-Eme1 endonuclease, and we found that codepletion of Mus81 and Wee1 abrogated the S phase delay. Importantly, Wee1 and Mus81 interact in vivo, suggesting direct regulation. Altogether, these results demonstrate a novel role of Wee1 in controlling Mus81 and DNA replication in human cells.
正确复制基因组并保护其完整性对于细胞存活至关重要。在一项研究 H2AX 磷酸化的高通量筛选中，我们发现 Wee1 是基因组稳定性的调节剂。下调 Wee1 不仅诱导了 H2AX 磷酸化，还触发了广泛的脱氧核糖核酸 (DNA) 损伤反应 (DDR)，并导致 DNA 复制受阻，使细胞积累在 S 期。Wee1 缺陷细胞的复制叉速度下降，表明 Wee1 参与了 DNA 复制。在短时间用羟基脲处理的细胞中抑制 Wee1 会增强 DDR，这表明 Wee1 专门保护停滞的复制叉的稳定性。值得注意的是，Wee1 耗竭诱导的 DDR 严重依赖于 Mus81-Eme1 内切酶，我们发现 Mus81 和 Wee1 的共耗竭消除了 S 期延迟。重要的是，Wee1 和 Mus81 在体内相互作用，表明存在直接调控。总之，这些结果表明 Wee1 在控制人类细胞中的 Mus81 和 DNA 复制方面发挥了新的作用。
