Depletion of Akt1 and Akt2 Impairs the Repair of Radiation-Induced DNA Double Strand Breaks via Homologous Recombination
Depletion of Akt1 and Akt2 Impairs the Repair of Radiation-Induced DNA Double Strand Breaks via Homologous Recombination
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DOI：
10.3390/ijms20246316
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发表时间：
2019-12-01
影响因子：
5.6
通讯作者：
Dittmann, Klaus
中科院分区：
文献类型：
--
作者：
Gol, Tahereh Mohammadian;Rodemann, H. Peter;Dittmann, Klaus
同源重组修复（HRR）、非同源末端连接（NHEJ）和替代性NHEJ是细胞处理DNA双链断裂（DNA-DSB）的主要途径，其功能在肿瘤细胞的辐射抗性中起重要作用。关于Akt在同源重组（HR）中的作用，即，Rad 51作为该途径的主要蛋白的调节。本研究旨在研究Akt亚型在HRR中的特异性参与。使用具有稳定的AKT敲除和siRNA介导的AKT敲除表型的HCT 116结肠癌细胞来研究Akt 1和Akt 2亚型在HR中的作用。结果清楚地表明，HCT 116 AKT 1-KO和AKT 2-KO细胞在照射后6 h与亲本细胞相比具有显著减少的Rad 51灶形成。Akt 1和Akt 2蛋白水平的消耗以及Akt激酶活性的抑制导致CENP-F阳性细胞中残留γ H2 AX的数量增加，主要代表S和G2期细胞。此外，使用DNA-PK和Rad 51拮抗剂抑制NHEJ和HR导致AKT 1和AKT 2敲除细胞比野生型细胞更强的放射敏感性。这些数据共同表明，Akt 1和Akt 2都通过HRR参与DSB修复。
Homologous recombination repair (HRR), non-homologous end-joining (NHEJ) and alternative NHEJ are major pathways that are utilized by cells for processing DNA double strand breaks (DNA-DSBs); their function plays an important role in the radiation resistance of tumor cells. Conflicting data exist regarding the role of Akt in homologous recombination (HR), i.e., the regulation of Rad51 as a major protein of this pathway. This study was designed to investigate the specific involvement of Akt isoforms in HRR. HCT116 colon cancer cells with stable AKT-knock-out and siRNA-mediated AKT-knockdown phenotypes were used to investigate the role of Akt1 and Akt2 isoforms in HR. The results clearly demonstrated that HCT116 AKT1-KO and AKT2-KO cells have a significantly reduced Rad51 foci formation 6 h post irradiation versus parental cells. Depletion of Akt1 and Akt2 protein levels as well as inhibition of Akt kinase activity resulted in an increased number of residual-gamma H2AX in CENP-F positive cells mainly representing the S and G2 phase cells. Furthermore, inhibition of NHEJ and HR using DNA-PK and Rad51 antagonists resulted in stronger radiosensitivity of AKT1 and AKT2 knockout cells versus wild type cells. These data collectively show that both Akt1 and Akt2 are involved in DSBs repair through HRR.