Department of Biochemistry and Molecular Biology, Department of Biomedical Sciences and Cancer Research Institute, Seoul National University College of Medicine, 103 Daehak-ro, Jongno-gu, Seoul, 03080, Korea.
Sci Rep. 2020 Sep 2;10(1):14455. doi: 10.1038/s41598-020-71522-9.
DNA double-strand breaks (DSB) are formed by various exogenous and endogenous factors and are repaired by homologous recombination and non-homologous end joining (NHEJ). DNA-dependent protein kinase (DNA-PK) is the principal enzyme for NHEJ. We explored the role and the underlying mechanism of cAMP signaling in the NHEJ repair of DSBs resulted from gamma ray irradiation to non-small cell lung cancer (NSLC) cells. Activated cAMP signaling by expression of an activated stimulatory GTP-binding protein or by pretreatment with isoproterenol and prostaglandin E2, delayed the repair of DSBs resulted from gamma ray irradiation, and the delaying effects depended on protein kinase A (PKA). Activated cAMP signaling suppressed XRCC4 and DNA ligase IV recruitment into DSB foci, and reduced phosphorylation at T2609 in DNA-PK catalytic subunit (DNA-PKcs) with a concomitant increase in phosphorylation at S2056 in PKA-dependent ways following gamma ray irradiation. cAMP signaling decreased phosphorylation of T2609 by protein phosphatase 2A-dependent inhibition of ATM. We conclude that cAMP signaling delays the repair of gamma ray-induced DNA DSBs in NSLC cells by inhibiting NHEJ via PKA-dependent pathways, and that cAMP signaling differentially modulates DNA-PKcs phosphorylation at S2056 and T2609, which might contribute to the inhibition of NHEJ in NSLC cells.
DNA 双链断裂 (DSB) 是由各种外源和内源因素形成的，并通过同源重组和非同源末端连接 (NHEJ) 进行修复。DNA 依赖性蛋白激酶 (DNA-PK) 是 NHEJ 的主要酶。我们研究了 cAMP 信号在非小细胞肺癌 (NSCLC) 细胞因γ射线照射而导致的 DSB 的 NHEJ 修复中的作用和潜在机制。通过表达激活的刺激 GTP 结合蛋白或用异丙肾上腺素和前列腺素 E2 预处理，激活的 cAMP 信号会延迟因γ射线照射而导致的 DSB 的修复，并且这种延迟作用取决于蛋白激酶 A (PKA)。激活的 cAMP 信号抑制 XRCC4 和 DNA 连接酶 IV 募集到 DSB 焦点，并减少 DNA-PK 催化亚基 (DNA-PKcs) 在 T2609 处的磷酸化，同时以 PKA 依赖性方式增加 S2056 处的磷酸化，这是在 γ 射线照射后发生的。cAMP 信号通过蛋白磷酸酶 2A 依赖性抑制 ATM 来减少 T2609 的磷酸化。我们得出结论，cAMP 信号通过 PKA 依赖性途径抑制 NHEJ，从而延迟 NSCLC 细胞中因 γ 射线诱导的 DNA DSB 的修复，并且 cAMP 信号在 S2056 和 T2609 处对 DNA-PKcs 的磷酸化产生不同的调节作用，这可能有助于抑制 NSCLC 细胞中的 NHEJ。
