目的：EGFR 是广泛分布于人体各组织细胞膜上的多功能糖蛋白，它可以通过磷酸化级联反应激活下游信号通路调控细胞行为，在肿瘤预防和治疗中的作用一直是研究的重点。然而有关于 EGFR 影响宫颈癌细胞辐射敏感性的研究较少且机制尚不明确。最近有研究显示，EGFR 可以通过 NRF2 信号通路发挥作用，但 EGFR 如何通过 NRF2 增加宫颈癌细胞的辐射敏感性还未阐明。本研究将探讨 EGFR 对宫颈癌细胞辐射敏感性的影响及 EGFR 在宫颈癌细胞受到电离辐射损伤后的 DNA 损伤响应机制中的作用。
方法：采用人宫颈癌 HeLa 细胞作为研究对象，使用小干扰 RNA 敲降细胞中EGFR 或 NRF2 的表达后进行 γ 射线照射；通过 CCK-8 实验观察 EGFR 和 NRF2对宫颈癌 HeLa 细胞增殖存活的影响；通过彗星实验、γ-H2AX 免疫荧光实验检测 EGFR 和 NRF2 对宫颈癌 HeLa 细胞 DNA 修复能力的影响；通过 Western Blot实验检测辐射后的 p-ATR (Ser1989)、p-CHK1 (Ser345)、HO-1 蛋白；通过流式细胞实验检测 EGFR 对宫颈癌 HeLa 细胞周期的影响；通过 RT-qPCR 实验检测EGFR、HO-1 mRNA 水平的变化。通过核质分离实验检测 EGFR 对 NRF2 核定位的影响；通过 Western Blot 实验检测 p-GSK-3β (Ser9)探究 EGFR 下游信号通路对 NRF2 出核的影响；另外还使用小干扰 RNA 敲降 KEAP1 或 MG132 抑制蛋白酶体处理宫颈癌 HeLa 细胞，观察 EGFR 对 NRF2 降解的影响。
结果：1、敲降 EGFR、NRF2 降低宫颈癌 HeLa 细胞增殖存活能力；
2、敲降 EGFR、NRF2 减弱宫颈癌 HeLa 细胞修复 DNA 损伤的能力；
3、敲降 EGFR、NRF2 阻碍了辐射后宫颈癌 HeLa 细胞的 G2/M 期细胞周期阻滞；
4、敲降 EGFR、NRF2 抑制了ATR/CHK1 介导的 DNA 损伤响应信号和抗氧化蛋白 HO-1 的表达；
5、敲降 EGFR 抑制辐射引起的 NRF2 核定位增加
6、EGFR 不通过 p-GSK-3β 信号通路促进 NRF2 出核；
7、EGFR 以不依赖于 KEAP1 介导的泛素蛋白酶体降解途径调控 NRF2 核分布；
结论：在宫颈癌 HeLa 细胞中，EGFR 以独立于 KEAP1 和 GSK-3β 的方式调控NRF2 核定位，下调抗氧化蛋白 HO-1 的表达，并通过 ATR-CHK1 信号通路减少辐射引起的细胞周期阻滞，降低宫颈癌 HeLa 细胞抗氧化能力和修复 DNA 损伤的能力，从而增加宫颈癌 HeLa 细胞的辐射敏感性。

Objective: EGFR is a multifunctional glycoprotein widely distributed on the cell membrane of various human tissues. It can regulate cell behavior by activating downstream signaling pathways through phosphorylation cascade reactions, and its role in tumor prevention and treatment has been a focus of research. However, there are fewer studies on EGFR affecting radiation sensitivity of cervical cancer cells and the mechanism remains unclear. Recently, it has been shown that EGFR can function through NRF2 signaling pathway. But how EGFR increases the radiation sensitivity of cervical cancer cells through NRF2 has not been elucidated. This study investigated the effect of EGFR on radiation sensitivity of cervical cancer cells and DNA damage response mechanism of EGFR in cervical cancer cells subjected to ionizing radiation damage. Methods: Small interfering RNA (siRNA) was used to knock down EGFR and NRF2 in HeLa cells。After γ-rays exposure, the cell survival and proliferation abilities of HeLa cells were observed by CCK-8 assay; comet assay and γ-H2AX immunofluorescence assay were performed to detect the effect of EGFR and NRF2 on DNA damage and repair ability of cervical cancer cells; The irradiated p-ATR (Ser1989), p-CHK1 (Ser345), HO-1 proteins were detected by Western blot assay; The effect of EGFR on cervical cancer cell cycle was detected by flow cytometry. EGFR, NRF2, HO-1 mRNA levels were measured by RT qPCR. Detection of p-GSK3β by Western blotting (Ser9) to explore the effect of EGFR downstream signaling pathway on NRF2 nucleation; Effect of EGFR on NRF2 degradation using siRNA knockdown of KEAP1 or MG132 inhibition of proteasome treatment of cervical cancer cells. Result: 1. Knocking down EGFR and NRF2 inhibited the cell survival and proliferative ability of cervical cancer HeLa cells; 2. Knocking down EGFR and NRF2 weakened the DNA damage repair ability of cervical cancer HeLa cells; 3. Knocking down EGFR and NRF2 hindered the G2/M phase cell cycle arrest of cervical cancer HeLa cells after radiation; 4. Knocking down EGFR and NRF2 inhibited ATR/CHK1 mediated DNA damage response signals and antioxidant protein HO-1 expression;  5. Knocking down EGFR suppressed radiation induced elevation of NRF2 nuclear localization 6. EGFR did not promote NRF2 out of nucleus through p-GSK-3 β; 7. EGFR regulates the nuclear distribution of NRF2 through a ubiquitin proteasome degradation pathway independent of KEAP1 mediation; Conclusions: In cervical cancer HeLa cells, EGFR regulated NRF2 nuclear entry in a KEAP1- independent manner, downregulated the expression of antioxidant proteins, and reduced radiation-induced cell cycle arrest through the ATR-CHK1 signaling pathway, which decreased the antioxidant capacity and DNA damage repair capacity of cervical cancer cells, thereby increasing the radiosensitivity of cervical cancer HeLa cells.