Dynamics of Cas10 Govern Discrimination between Self and Non-self in Type III CRISPR-Cas Immunity

Adaptive immune systems must accurately distinguish between self and non-self in order to defend against invading pathogens while avoiding autoimmunity. Type III CRISPR-Cas systems employ guide RNA to recognize complementary RNA targets, which triggers the degradation of both the invader’s transcripts and their template DNA. These systems can broadly eliminate foreign targets with multiple mutations but circumvent damage to the host genome. To explore the molecular basis for these features, we use single-molecule fluorescence microscopy to study the interaction between a type III-A ribonucleoprotein complex and various RNA substrates. We find that Cas10—the DNase effector of the complex—displays rapid conformational fluctuations on foreign RNA targets, but is locked in a static configuration on self RNA. Target mutations differentially modulate Cas10 dynamics and tune the CRISPR interference activity in vivo. These findings highlight the central role of the internal dynamics of CRISPR-Cas complexes in self versus non-self discrimination and target specificity. [Display omitted] •Self RNA locks Cas10 in a static inactive configuration, thus avoiding autoimmunity•Non-self RNA triggers fast conformational fluctuations within Cas10 and activates it•Target mismatches differentially modulate the conformational dynamics of Cas10•Internal flexibility of the effector complex enables tunable CRISPR immune response Type III CRISPR-Cas systems employ an RNA-DNA dual-targeting mechanism to eliminate invading pathogens. Using single-molecule fluorescence spectroscopy, Wang et al. show that the conformational dynamics of Cas10—the signature subunit of type III effector complexes—control the discrimination between self and non-self elements and the strength of CRISPR immunity.