Structure of the IscB-ωRNA ribonucleoprotein complex, the likely ancestor of CRISPR-Cas9

Transposon-encoded IscB family proteins are RNA-guided nucleases in the OMEGA (obligate mobile element-guided activity) system, and likely ancestors of the RNA-guided nuclease Cas9 in the type II CRISPR-Cas adaptive immune system. IscB associates with its cognate omega RNA to form a ribonucleoprotein complex that cleaves double-stranded DNA targets complementary to an omega RNA guide segment. Although IscB shares the RuvC and HNH endonuclease domains with Cas9, it is much smaller than Cas9, mainly due to the lack of the alpha-helical nucleic-acid recognition lobe. Here, we report the cryo-electron microscopy structure of an IscB protein from the human gut metagenome (OgeuIscB) in complex with its cognate omega RNA and a target DNA, at 2.6-angstrom resolution. This high-resolution structure reveals the detailed architecture of the IscB-omega RNA ribonucleoprotein complex, and shows how the small IscB protein assembles with the omega RNA and mediates RNA-guided DNA cleavage. The large omega RNA scaffold structurally and functionally compensates for the recognition lobe of Cas9, and participates in the recognition of the guide RNA-target DNA heteroduplex. These findings provide insights into the mechanism of the programmable DNA cleavage by the IscB-omega RNA complex and the evolution of the type II CRISPR-Cas9 effector complexes.

Automated summary

This study reports the structure of the IscB protein in complex with its cognate omega RNA and a target DNA from the human gut. The structure reveals the detailed architecture of the IscB-omega RNA ribonucleoprotein complex and shows how IscB assembles with the omega RNA to mediate RNA-guided DNA cleavage. The large omega RNA scaffold compensates for the lack of the recognition lobe in Cas9 and participates in the recognition of the guide RNA-target DNA heteroduplex.