Prespacers formed during primed adaptation associate with the Cas1-Cas2 adaptation complex and the Cas3 interference nuclease-helicase.

For Type I CRISPR-Cas systems, a mode of CRISPR adaptation named priming has been described. Priming allows specific and highly efficient acquisition of new spacers from DNA recognized (primed) by the Cascade-crRNA (CRISPR RNA) effector complex. Recognition of the priming protospacer by Cascade-crRNA serves as a signal for engaging the Cas3 nuclease-helicase required for both interference and primed adaptation, suggesting the existence of a primed adaptation complex (PAC) containing the Cas1-Cas2 adaptation integrase and Cas3. To detect this complex in vivo, we here performed chromatin immunoprecipitation with Cas3-specific and Cas1-specific antibodies using cells undergoing primed adaptation. We found that prespacers are bound by both Cas1 (presumably, as part of the Cas1-Cas2 integrase) and Cas3, implying direct physical association of the interference and adaptation machineries as part of PAC.