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The cGAS-Ku80 complex regulates the balance between two end joining subpathways

Cell Death & Differentiation (2024)Cite this article

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Abstract

As the major DNA sensor that activates the STING-TBK1 signaling cascade, cGAS is mainly present in the cytosol. A number of recent reports have indicated that cGAS also plays critical roles in the nucleus. Our previous work demonstrated for the first time that cGAS is translocated to the nucleus upon the occurrence of DNA damage and inhibits homologous recombination (HR), one of the two major pathways of DNA double strand break (DSB) repair. However, whether nuclear cGAS regulates the other DSB repair pathway, nonhomologous end joining (NHEJ), which can be further divided into the less error-prone canonical NHEJ (c-NHEJ) and more mutagenic alternative NHEJ (alt-NHEJ) subpathways, has not been characterized. Here, we demonstrated that cGAS tipped the balance of the two NHEJ subpathways toward c-NHEJ. Mechanistically, the cGAS-Ku80 complex enhanced the interaction between DNA-PKcs and the deubiquitinase USP7 to improve DNA-PKcs protein stability, thereby promoting c-NHEJ. In contrast, the cGAS-Ku80 complex suppressed alt-NHEJ by directly binding to the promoter of Polθ to suppress its transcription. Together, these findings reveal a novel function of nuclear cGAS in regulating DSB repair, suggesting that the presence of cGAS in the nucleus is also important in the maintenance of genome integrity.

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Fig. 1: cGAS promotes c-NHEJ and suppresses alt-NHEJ.
Fig. 2: cGAS increases DNA-PKcs protein stability.
Fig. 3: cGAS promotes USP7-mediated DNA-PKcs deubiquitination and stabilization.
Fig. 4: cGAS interacts with Ku80 to promote USP7-mediated stabilization of DNA-PKcs.
Fig. 5: cGAS inhibits the transcription of Polθ.
Fig. 6: cGAS inhibits alt-NHEJ by suppressing Polθ transcription in a Ku80-dependent manner.

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All data is contained within the manuscript and/or supplementary files. The original western blot images are provided in the Supplementary File.

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Acknowledgements

We thank Dr. Jiemin Wong and Jialun Li (East China Normal University, Shanghai, China) for generously providing the Flag-USP7 and Flag-USP7m plasmids. We thank Dr. Haiying Wang (Peking University Health Science Center, Beijing, China) for providing the GST-USP7, GST-USP7-TRAF, GST-USP7-CAT, GST-USP7-UBL and HA-USP7 plasmids. We thank Dr. Yanhui Xu and Xiaotong Yin (Fudan University, Shanghai, China) for their generous provision of DNA-PKcs proteins, KU70/80 proteins and P53 proteins.

Funding

This work was supported by the National Key R&D Program of China (grant numbers 2021YFA1102003 and 2022YFA1103703 to Z.M.) and the National Natural Science Foundation of China (grant numbers 32171288 to Y.J., 82225017, 32270750 and 82071565 to Z.M., and 82101634 to H.Z.), and the Shanghai Sailing Program (21YF1435900 to H.Z.).

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  1. These authors contributed equally: Haiping Zhang, Lijun Jiang.

Authors and Affiliations

  1. Shanghai Key Laboratory of Maternal Fetal Medicine, Clinical and Translational Research Center of Shanghai First Maternity and Infant Hospital, Frontier Science Center for Stem Cell Research, School of Life Sciences and Technology, Tongji University, Shanghai, China

    Haiping Zhang, Lijun Jiang, Zhen Qian, Ying Jiang & Zhiyong Mao

  2. Department of Gynecology, Shanghai First Maternity and Infant Hospital, School of Medicine, Tongji University, Shanghai, China

    Xinyi Du & Guizhu Wu

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HZ, ZM, YJ and GW conceived the project, designed the experiments, and wrote the paper. HZ and LJ performed most of the experiments and analyzed the data. ZQ and XD assisted with the acquisition of data.

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Correspondence to Guizhu Wu, Ying Jiang or Zhiyong Mao.

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Zhang, H., Jiang, L., Du, X. et al. The cGAS-Ku80 complex regulates the balance between two end joining subpathways. Cell Death Differ (2024). https://doi.org/10.1038/s41418-024-01296-4

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