He Yuqiu, Wang Qing, Zhang Qingqing, Wang Yifei, Jiang Yuqian, Zhao Qiu, Liu Xiaoqing, Wang Fuan
College of Chemistry and Molecular Sciences, Wuhan University, Wuhan, 430072, China.
Department of Gastroenterology, Zhongnan Hospital of Wuhan University, Hubei Provincial Clinical Research Center for Intestinal and Colorectal Diseases, Hubei Key Laboratory of Intestinal and Colorectal Diseases, Wuhan, 430071, China.
Small Methods. 2025 Jun;9(6):e2401160. doi: 10.1002/smtd.202401160. Epub 2024 Sep 19.
The on-demand gene regulation is crucial for extensively exploring specific gene functions and developing personalized gene therapeutics, which shows great promise in precision medicines. Although some nucleic acid-based gene regulatory tools (antisense oligonucleotides and small interfering RNAs) are devised for achieving on-demand activation, the introduction of chemical modifications may cause undesired side effects, thereby impairing the gene regulatory efficacy. Herein, a methyl-engineered DNAzyme (MeDz) is developed for the visualization of endogenous alkyltransferase (AGT) and the simultaneous self-sufficiently on-demand gene regulation. The catalytic activity of DNAzyme can be efficiently blocked by O-methylguanine (OMeG) modification and specifically restored via the AGT-mediated DNA-repairing pathway. This simply designed MeDz is demonstrated to reveal AGT of varying expression levels in different cells, opening the possibility to explore the AGT-related biological processes. Moreover, the AGT-guided MeDz exhibits cell-selective regulation on the human early growth response-1 (EGR-1) gene, with efficient gene repression in breast cancer cells and low effectiveness in normal cells. The proposed MeDz offers an attractive strategy for on-demand gene regulation, displaying great potential in biomedical applications.
按需基因调控对于广泛探索特定基因功能和开发个性化基因疗法至关重要，这在精准医学中显示出巨大潜力。尽管一些基于核酸的基因调控工具（反义寡核苷酸和小干扰RNA）被设计用于实现按需激活，但化学修饰的引入可能会导致不良副作用，从而损害基因调控效果。在此，开发了一种甲基工程化脱氧核酶（MeDz），用于可视化内源性烷基转移酶（AGT）并同时实现自给自足的按需基因调控。脱氧核酶的催化活性可被O-甲基鸟嘌呤（OMeG）修饰有效阻断，并通过AGT介导的DNA修复途径特异性恢复。这种设计简单的MeDz被证明可以揭示不同细胞中不同表达水平的AGT，为探索与AGT相关的生物学过程开辟了可能性。此外，AGT引导的MeDz对人类早期生长反应-1（EGR-1）基因表现出细胞选择性调控，在乳腺癌细胞中具有高效的基因抑制作用，而在正常细胞中效果较低。所提出的MeDz为按需基因调控提供了一种有吸引力的策略，在生物医学应用中显示出巨大潜力。
