Cas13 has demonstrated unique and broad utility in RNA editing, nucleic acid detection, and disease diagnosis; however, a constantly active Cas enzyme may induce unwanted effects. Bacteriophage- or prophage-region-encoded anti-CRISPR (acr) gene molecules provide the potential to control targeting specificity and potency to allow for optimal RNA editing and nucleic acid detection by spatiotemporally modulating endonuclease activities. Using integrated approaches to screen acrVI candidates and evaluate their effects on Cas13 function, we discovered a series of acrVIA1-7 genes that block the activities of Cas13a. These VI-A CRISPR inhibitors substantially attenuate RNA targeting and editing by Cas13a in human cells. Strikingly, type VI-A anti-CRISPRs (AcrVIAs) also significantly muffle the single-nucleic-acid editing ability of the dCas13a RNA-editing system. Mechanistically, AcrVIA1, -4, -5, and -6 bind LwaCas13a, while AcrVIA2 and -3 can only bind the LwaCas13-crRNA (CRISPR RNA) complex. These identified acr molecules may enable precise RNA editing in Cas13-based application and study of phage-bacterium interaction.

Cas13在RNA编辑，核酸检测和疾病诊断中显示出独特而广泛的用途。但是，持续活跃的Cas酶可能会引起不良影响。噬菌体或噬菌体区域编码的抗CRISPR（acr）基因分子提供了控制靶向特异性和潜能的潜力，从而可以通过时空调节核酸内切酶的活性实现最佳的RNA编辑和核酸检测。使用集成方法筛选acrVI候选物并评估其对Cas13功能的影响，我们发现了一系列acrVIA1-7阻断Cas13a活性的基因。这些VI-A CRISPR抑制剂可大大减弱Cas13a在人细胞中的RNA靶向和编辑。引人注目的是，VI-A型抗CRISPRs（AcrVIA）也大大抑制了dCas13a RNA编辑系统的单核酸编辑能力。从机理上讲，AcrVIA1，-4，-5和-6结合LwaCas13a，而AcrVIA2和-3只能结合LwaCas13-crRNA（CRISPR RNA）复合体。这些鉴定的acr分子可以在基于Cas13的应用和噬菌体-细菌相互作用的研究中实现精确的RNA编辑。