Core Viewpoint - The article discusses the CRISPR-Cas system as a significant tool in gene editing and disease treatment, highlighting its immune mechanisms and the role of Cas9 in regulating spacer acquisition and immune memory formation [3][8][15]. Group 1: CRISPR-Cas System Overview - The CRISPR-Cas system is a bacterial immune mechanism that utilizes "molecular memory" and "precise cutting" to defend against foreign invaders [3]. - It consists of two main components: the CRISPR array, which serves as a memory bank, and Cas proteins, which are responsible for recognizing and cutting foreign DNA [3][4]. Group 2: Immune Process Stages - The immune process mediated by CRISPR-Cas can be divided into three stages: Adaptation, Expression, and Interference [4]. - During the Adaptation stage, the Cas1-Cas2 complex integrates foreign DNA sequences into the CRISPR array [4]. - The Expression stage involves the transcription of the CRISPR array into pre-crRNA, which is then processed into mature crRNA [4]. - In the Interference stage, Cas proteins form an effector complex with guide RNA to identify and cleave complementary foreign DNA [4]. Group 3: Research Findings on Cas9 - Recent studies reveal that the II-C type system has a unique molecular regulation mechanism during the immune adaptation phase, where apoCas9 significantly enhances spacer acquisition efficiency [8][10]. - The research indicates that apoCas9 acts as both a sensor for crRNA abundance and a regulator of spacer acquisition efficiency, challenging previous beliefs about its inactivity [8][15]. - The study also highlights the importance of the NUC lobe in driving efficient spacer acquisition, while the REC lobe modulates this process through RNA interactions [12][14]. Group 4: Implications and Future Directions - The findings provide insights into how bacteria can quickly replenish their immune memory after CRISPR array contraction, emphasizing the dynamic regulation of immune memory depth [15]. - The research lays a theoretical foundation for future applications in CRISPR-based immune adaptation machines, molecular recording, lineage tracing, and anti-phage engineering [17].