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 approximately 20S and approximately 35-40S and present the three-dimensional structures of both complexes by electron microscopy. The approximately 35-40S complexes consist of a platform density packed against a semispherical element. The approximately 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 approximately 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.
Golas
Monika M
MM
Research Group of 3D Electron Cryomicroscopy, Max Planck Institute for Biophysical Chemistry, Göttingen, Germany.
Böhm
Cordula
C
Sander
Bjoern
B
Effenberger
Kerstin
K
Brecht
Michael
M
Stark
Holger
H
Göringer
H Ulrich
HU
D000818:Animals; D020285:Cryoelectron Microscopy; D008954:Models, Biological; D008958:Models, Molecular; D015800:Protozoan Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000648:ultrastructure; D017393:RNA Editing; D016053:RNA, Protozoan / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism* / Q000648:ultrastructure; D014345:Trypanosoma / Q000378:metabolism* / Q000648:ultrastructure; D014461:Ultracentrifugation
D015800:Protozoan Proteins; D016053:RNA, Protozoan