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.