Snapshots of the RNA editing machine in trypanosomes captured at different assembly stages in vivo

Mitochondrial pre-messenger RNAs in kinetoplastid protozoa are substrates of uridylate-specific RNA editing. RNA editing converts non-functional pre-mRNAs into translatable molecules and can generate protein diversity by alternative editing. Although several editing complexes have been described, their structure and relationship is unknown. Here, we report the isolation of functionally active RNA editing complexes by a multistep purification procedure. We show that the endogenous isolates contain two subpopulations of similar to 20S and similar to 35-40S and present the three-dimensional structures of both complexes by electron microscopy. The similar to 35-40S complexes consist of a platform density packed against a semispherical element. The similar to 20S complexes are composed of two subdomains connected by an interface. The two particles are structurally related, and we show that RNA binding is a main determinant for the interconversion of the two complexes. The similar to 20S editosomes contain an RNA-binding site, which binds gRNA, pre-mRNA and gRNA/pre-mRNA hybrid molecules with nanomolar affinity. Variability analysis indicates that subsets of complexes lack or possess additional domains, suggesting binding sites for components. Together, a picture of the RNA editing machinery is provided.