A target expression threshold dictates invader defense and autoimmunity by CRISPR-Cas13

Immune systems must recognize and clear foreign invaders without eliciting autoimmunity. CRISPR-Cas immune systems in prokaryotes manage this task by following two criteria: extensive guide:target complementarity and a defined target-flanking motif. Here we report an additional requirement for RNA-targeting CRISPR-Cas13 systems: expression of the target transcript exceeding a threshold. This finding is based on targeting endogenous non-essential transcripts, which rarely elicited dormancy through collateral RNA degradation. Instead, eliciting dormancy required over-expressing targeted transcripts above a threshold. A genome-wide screen confirmed target expression levels as the principal determinant of cytotoxic autoimmunity and revealed that the threshold shifts with the guide:target pair. This expression threshold ensured defense against a lytic bacteriophage yet allowed tolerance of a targeted beneficial gene expressed from an invading plasmid. These findings establish target expression levels as a third criterion for immune defense by RNA-targeting CRISPR-Cas systems, buffering against autoimmunity and distinguishing pathogenic and benign invaders.

免疫系统需识别并清除外来入侵者，同时避免引发自身免疫反应。原核生物中的CRISPR-Cas免疫系统通过遵循两项标准来完成这一任务：广泛的引导序列与靶序列互补性以及明确的靶序列旁侧基序。本研究报告了RNA靶向CRISPR-Cas13系统的一个额外要求：靶转录本的表达量超过一定阈值。这一发现基于靶向内源非必需转录本，这些转录本很少通过旁侧RNA降解引起休眠。相反，引发休眠需要超过阈值表达靶向转录本。全基因组筛选证实靶转录本的表达水平是细胞毒性自身免疫的主要决定因素，并揭示了阈值随着引导序列与靶序列对的变化而变化。这一表达阈值确保了对裂解性噬菌体的防御，同时允许对来自入侵性质粒表达的靶向有益基因的耐受。这些发现将靶转录本的表达水平确立为RNA靶向CRISPR-Cas系统免疫防御的第三项标准，在自身免疫的缓冲作用中发挥作用，并区分病原性和良性入侵者。