An essential role of KREPB4 in RNA editing and structural integrity of the editosome in Trypanosoma brucei

Abstract

RNA editing in the sleeping sickness parasite Trypanosoma brucei remodels mitochondrial transcripts by the addition and deletion of uridylates as specified by guide RNAs. Editing is catalyzed by at least three distinct ∼20S multiprotein editosomes, all of which contain KREPB4, a protein with RNase III and Pumilio motifs. RNAi repression of KREPB4 expression in procyclic forms (PFs) strongly inhibited growth and in vivo RNA editing, greatly diminished the abundance of 20S editosomes, reduced cellular levels of editosome proteins, and generated ∼5-10S editosome subcomplexes. Editing TUTase, exoUase, and RNA ligase activities were largely shifted from ∼20S to ∼5-10S fractions of cellular lysates. Insertion and deletion endonuclease activities in ∼20S fractions were strongly reduced upon KREPB4 repression but were not detected in the 5-10S subcomplex fraction. Abundance of MRP1 and RBP16 proteins, which appear to be involved in RNA processing but are not components of the 20S editosome, was unaltered upon KREPB4 repression. These data suggest that KREPB4 is important for the structural integrity of ∼20S editosomes, editing endonuclease activity, and the viability of PF T. brucei cells. Published by Cold Spring Harbor Laboratory Press. Copyright © 2007 RNA Society.