CRISPR-Cas systems provide prokaryotes with acquired immunity against viruses and plasmids, but how these systems are regulated to prevent autoimmunity is poorly understood. Here, we show that in the S. pyogenes CRISPR-Cas system, a long-form transactivating CRISPR RNA (tracr-L) folds into a natural single guide that directs Cas9 to transcriptionally repress its own promoter (P_cas). Further, we demonstrate that P_cas serves as a critical regulatory node. De-repression causes a dramatic 3,000-fold increase in immunization rates against viruses; however, heightened immunity comes at the cost of increased autoimmune toxicity. Using bioinformatic analyses, we provide evidence that tracrRNA-mediated autoregulation is widespread in type II-A CRISPR-Cas systems. Collectively, we unveil a new paradigm for the intrinsic regulation of CRISPR-Cas systems by natural single guides, which may facilitate the frequent horizontal transfer of these systems into new hosts that have not yet evolved their own regulatory strategies.

CRISPR-Cas 系统为原核生物提供了针对病毒和质粒的获得性免疫，但如何调节这些系统以防止自身免疫，我们知之甚少。在这里，我们展示了在化脓性链球菌CRISPR-Cas 系统中，长形式的反式激活 CRISPR RNA (tracr-L)折叠成一个天然的单一向导，指导 Cas9 转录抑制其自身的启动子 (P _cas )。此外，我们证明 P _cas作为一个关键的监管节点。去抑制导致对病毒的免疫率急剧增加 3,000 倍；然而，增强免疫力是以增加自身免疫毒性为代价的。使用生物信息学分析，我们提供证据表明tracrRNA介导的自动调节在 II-A 型 CRISPR-Cas 系统中很普遍。总的来说，我们揭示了一种通过自然单一指南对 CRISPR-Cas 系统进行内在调控的新范式，这可能有助于将这些系统频繁地横向转移到尚未进化出自己的调控策略的新宿主中。