在基于CRISPR的转录激活系统中激活体表达的细胞毒性
Ziyan Liang1, Aakaanksha Maddineni1, Jesus A Ortega1
1Department of Microbiology-Immunology, Northwestern University, Feinberg School of Medicine, Chicago, IL, USA.
Nature communications
|August 28, 2025
概括
常见的基于CRISPR的转录激活系统 (CRISPRa) 显示出显著的细胞毒性. 这种与激活域相关的细胞毒性限制了CRISPRa
科学领域:
- 分子生物学
- 基因编辑技术
- 细胞毒理学
背景情况:
- 基于CRISPR的转录激活 (CRISPRa) 对研究和临床应用具有重大潜力.
- CRISPRa系统使用转录激活剂来调节基因表达.
- 协同激活媒介 (SAM) CRISPRa系统是一个广泛使用的方法.
研究的目的:
- 研究常用的CRISPRa系统,特别是SAM系统的细胞毒性.
- 确定导致观察到的毒性的特定成分.
- 评估这种毒性对CRISPRa应用的影响.
主要方法:
- 对表达特定激活域 (AD) 和乙转移酶核心域的CRISPRa载体的评估.
- 在生产细胞中评估晶状病毒标位.
- 在转化目标细胞中分析细胞活力.
- 使用可诱导的透视病毒载体来探索激活器表达窗口.
主要成果:
- 表达p65和HSF1激活域的CRISPRa载体具有明显的细胞毒性.
- 强大的转录激活剂的高水平表达导致目标细胞的低位和细胞死亡.
- 使用可诱导系统没有发现基于SAM的CRISPRa安全窗口,这表明固有毒性.
结论:
- 目前基于SAM的CRISPRa系统的细胞毒性对其在生物医学研究中的广泛采用构成了重大障碍.
- 这种毒性可以在遗传选中引入混的选择压力.
- 未来的CRISPRa技术开发必须优先考虑激活效率和减轻激活剂毒性.
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