近日，中国科学院化学研究所程靓团队实现了用于光控CRISPR/Cas编辑的功能可调星形多价crRNAs。该研究于2025年4月14日发表在《德国应用化学》杂志上。

基于簇状规则间隔短回文重复序列/CRISPR相关（CRISPR/Cas）的基因组编辑因其精确性、简单性和多功能性而显著推进了基因工程。然而，实现精确的空间和时间控制仍然具有挑战性，限制了治疗和研究应用。研究组介绍了一类新的星形多价crRNAs，用于CRISPR/Cas9和Cas12a编辑系统的精确时空控制。这些crRNAs是通过单位点化学修饰合成的，可以有效纯化。

通过整合不同的光响应化学连接，研究组在特定波长的照射下实现了crRNA活性的选择性激活，从而能够同时对多个遗传靶点进行正交调节。该方法在体外证明了强大的关断开关能力，其特征是泄漏最小，激活快速。重要的是，该方法还被证明对哺乳动物细胞体内的时间控制基因编辑非常有效，在短暂的光活化后实现了相当高的编辑效率。由于其与靶序列无关的单位点修饰设计，该策略可以作为多种CRISPR/Cas系统的通用解决方案，消除繁琐的优化过程。结合长波长响应和可逆连接体的未来进展有望进一步增强组织渗透和控制，显著扩大这种方法在生物研究和治疗干预中的适用性和影响。

附：英文原文

Title: Functionally Tunable Star-Shaped Multivalent crRNAs for Photocontrol CRISPR/Cas Editing

Author: Wen-Da Chen, Li Liu, Liang Cheng

Issue&Volume: 2025-04-14

Abstract: Clustered regularly interspaced shortpalindromic repeats/CRISPR-associated (CRISPR/Cas)-based genome editing has significantly advanced genetic engineering due to its precision, simplicity, and versatility.  However, achieving precise spatial and temporal control remains challenging, restricting therapeutic and research applications.  Herein, we introduce a novel class of star-shaped, multivalent crRNAs engineered for precise spatiotemporal control of CRISPR/Cas9 and Cas12a editing systems.  These crRNAs are synthesized via single-site chemical modification and can be efficiently purified.  By integrating distinct photo-responsive chemical linkages, we achieved selective activation of crRNA activity upon irradiation with specific wavelengths, enabling orthogonal regulation of multiple genetic targets simultaneously.  This method demonstrated robust OFF-ON switching capabilities in vitro, characterized by minimal leakage and rapid activation.  Importantly, the approach also proved highly effective for temporally controlled gene editing in mammalian cells in vivo, achieving considerable editing efficiency following brief photoactivation.  Due to its target sequence-independent, single-site modification design, this strategy may serve as a universal solution for diverse CRISPR/Cas systems, eliminating cumbersome optimization processes.  Future advancements incorporating long-wavelength responsive and reversible linkers promise further enhancement of tissue penetration and control, significantly broadening the applicability and impact of this approach in biological research and therapeutic interventions.

DOI: 10.1002/anie.202506527

Source: https://onlinelibrary.wiley.com/doi/10.1002/anie.202506527