Native mRNA editing complexes from Trypanosoma brucei mitochondria.

Abstract

The aim of this study was to identify multicomponent complexes involved in kinetoplastid mitochondrial mRNA editing. Mitochondrial extracts from Trypanosoma brucei were fractionated on 10–30% glycerol gradients and assayed for RNAs and activities potentially involved in editing, including pre‐edited mRNA, guide RNA (gRNA), endonuclease, terminal uridylyltransferase (TUTase), RNA ligase and gRNA‐mRNA chimera‐forming activities. These experiments suggest that two distinct editing complexes exist. Complex I (19S) consists of gRNA, TUTase, RNA ligase and chimera‐forming activity. Complex II (35–40S) is composed of gRNA, preedited mRNA, RNA ligase and chimera‐forming activity. These studies provide the first evidence that editing occurs in a multicomponent complex. The possible roles of complex I, complex II and RNA ligase in editing are discussed.

Trypanosoma brucei gBP21

“…The ability of gBP21 to cross-link to added synthetic gRNAs within an E. coli cell extract as well as in a purified form suggests, that the binding of gRNAs to gBP21 is not dependent on the interaction of other mitochondrial proteins. Although we cannot exclude a stimulatory effect by other polypeptides, this might indicate that gBP21 is an early assembly component to gRNAs which form several high molecular weight RNP complexes in T. brucei mitochondria (8,18). In contrast, it is equally possible that gBP21 by binding to gRNAs prevents the base pairing interaction to the pre-mRNAs.…”

“…3) and RNA editing in vitro requires mitochondrial protein extracts (4 -7). Associated with the RNP complexes are various enzymatic activities such as endonuclease (8,9), RNA ligase (8 -10), RNA helicase (11,12), and terminal uridylyltransferase (8,12,13), which have been suggested to be catalytic components of different steps of the RNA editing process. Corell et al (12) determined an apparent S value of 20 S for a mitochondrial uridylate deletion activity.…”

RNA editing in kinetoplastid protozoa

“…In the cleavage-ligation (CL) model, Blum et al (15) proposed a series of steps that are catalyzed by the protein enzymes endoribonuclease, TUTase, and RNA ligase. These activities have been demonstrated in the mt of kinetoplastids (7,41,71). Enzyme-catalyzed steps are also part of the cleavage-ligation/chimera (CL-C) model proposed by Sollner-Webb (99), and gRNA/mRNA chimeric molecules (chimeras), which have been detected in cellular RNA, are purported editing intermediates.…”

RNA editing in kinetoplastid parasites: What to do with U

“…It consists of a single network of thousands of catenated circular DNAs of two size classes: minicircles of 0.9-2.5 kb (depending on the species), which are heterogeneous in sequence and account for the major portion of the network, and 20-50 homoplasmic maxicircles with an interspecies size variation of [23][24][25][26][27][28][29][30][31][32][33][34][35][36][37][38][39][40]. Mt gene expression appears to be a collaborative effort by both maxicircles and minicircles (see Box 3 and Fig.…”