Wang Feifei, Gui Wenli, Rong Mengtao, Zhang Liang, Wu Jiajing, Li Juan, Wang Renqing, Gouttia Odjo G, Wang Ling, Yang Xingyuan, Peng Aimin
Institute of Health Sciences and Technology, Institutes of Physical Sciences and Information Technology, Anhui University, Hefei, Anhui, China.
Institute of Health Sciences and Technology, Institutes of Physical Sciences and Information Technology, Anhui University, Hefei, Anhui, China.
J Biol Chem. 2025 May 4;301(6):110174. doi: 10.1016/j.jbc.2025.110174.
Nonhomologous end joining (NHEJ) is a pivotal mechanism in the repair of DNA double-strand breaks. Central to NHEJ is the DNA-dependent protein kinase (DNA-PK) complex, comprising the KU heterodimer and the catalytic subunit, DNA-PKcs. In this study, we characterize Thymocyte Selection-Associated High-Mobility Group Box Family Member 4 (TOX4) as a factor recruited to both laser-induced DNA damage and endonuclease-induced DNA double-strand breaks. Depletion of TOX4 leads to accumulation of DNA damage, which is epistatic to DNA-PKcs. Consistently, TOX4 depletion substantially reduces NHEJ efficiency measured using both intrachromosomal and extrachromosomal repair assays. Our proteomic and biochemical analyses reveal TOX4 association with DNA-PK that is required for DNA-PKcs activation. Furthermore, we show that TOX4 coordinates with phosphatase 1 nuclear-targeting subunit in NHEJ. Phosphatase 1 nuclear-targeting subunit, previously shown to protect DNA-PKcs phosphorylation from protein phosphatase 1-mediated dephosphorylation, binds DNA-PK in a TOX4-dependent manner. In line with its role in DNA repair, TOX4 emerges as a promising target for anticancer drug development, and its targeting enhances tumor cell sensitivity to DNA damage in head and neck cancer and other malignancies.
非同源末端连接（NHEJ）是DNA双链断裂修复中的关键机制。NHEJ的核心是DNA依赖性蛋白激酶（DNA-PK）复合物，它由KU异二聚体和催化亚基DNA-PKcs组成。在本研究中，我们将胸腺细胞选择相关高迁移率族框蛋白4（TOX4）鉴定为一种被募集到激光诱导的DNA损伤和核酸内切酶诱导的DNA双链断裂处的因子。TOX4的缺失导致DNA损伤积累，这在基因上位关系上对DNA-PKcs有影响。一致地，使用染色体内部和染色体外修复试验测量，TOX4的缺失显著降低了NHEJ效率。我们的蛋白质组学和生化分析揭示了TOX4与DNA-PK的关联，这是DNA-PKcs激活所必需的。此外，我们表明TOX4在NHEJ中与磷酸酶1核靶向亚基协同作用。磷酸酶1核靶向亚基先前已被证明可保护DNA-PKcs磷酸化免受蛋白磷酸酶1介导的去磷酸化作用，它以TOX4依赖的方式与DNA-PK结合。与其在DNA修复中的作用一致，TOX4成为抗癌药物开发的一个有前景的靶点，靶向TOX4可增强头颈癌和其他恶性肿瘤中肿瘤细胞对DNA损伤的敏感性。
