Uridine insertion/deletion RNA editing in trypanosome mitochondria: A complex business

Simpson, Larry; Sbicego, Sandro; Aphasizhev, Ruslan

doi:10.1261/rna.2178403

Cited by 154 publications

(103 citation statements)

References 74 publications

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…editing ͉ electron microscopy ͉ kinetoplast ͉ Leishmania ͉ trypanosome U ridine (U) insertion/deletion RNA editing is a posttranscriptional process in mitochondria of kinetoplastid protists that involves the modification of mRNA transcripts of mitochondrial cryptogenes by precise insertion and deletion of U residues to create translatable sequences (1)(2)(3). We previously proposed a model for the mechanism of this editing process involving a cleavage, addition or deletion of U's and ligation progressing in a 3Ј to 5Ј polarity (4), and this model has been since experimentally validated in almost every detail (5)(6)(7)(8)(9)(10).…”

mentioning

confidence: 99%

Structure of the core editing complex (L-complex) involved in uridine insertion/deletion RNA editing in trypanosomatid mitochondria

Li¹,

Hui

et al. 2009

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

View full text Add to dashboard Cite

Uridine insertion/deletion RNA editing is a unique form of posttranscriptional RNA processing that occurs in mitochondria of kinetoplastid protists. We have carried out 3D structural analyses of the core editing complex or ''L (ligase)-complex'' from Leishmania tarentolae mitochondria isolated by the tandem affinity purification procedure (TAP). The purified material, sedimented at 20 -25S, migrated in a blue native gel at 1 MDa and exhibited both precleaved and full-cycle gRNA-mediated U-insertion and U-deletion in vitro activities. The purified L-complex was analyzed by electron tomography to determine the extent of heterogeneity. Three-dimensional structural comparisons of individual particles in the tomograms revealed that a majority of the complexes have a similar shape of a slender triangle. An independent single-particle reconstruction, using a featureless Gaussian ball as the initial model, converged to a similar triangular structure. Another singleparticle reconstruction, using the averaged tomography structure as the initial model, yielded a similar structure. The REL1 ligase was localized on the model to the base of the apex by decoration with REL1-specific IgG. This structure should prove useful for a detailed analysis of the editing reaction.editing ͉ electron microscopy ͉ kinetoplast ͉ Leishmania ͉ trypanosome U ridine (U) insertion/deletion RNA editing is a posttranscriptional process in mitochondria of kinetoplastid protists that involves the modification of mRNA transcripts of mitochondrial cryptogenes by precise insertion and deletion of U residues to create translatable sequences (1-3). We previously proposed a model for the mechanism of this editing process involving a cleavage, addition or deletion of U's and ligation progressing in a 3Ј to 5Ј polarity (4), and this model has been since experimentally validated in almost every detail (5-10).Editing involves multiple multiprotein complexes associated by RNA linkers. The core editing complex, known as the L-complex, ''20S complex,'' ''editosome,'' or ''ϳ20S editosome,'' was detected by following the sedimentation and gel filtration of 2 adenylatable RNA ligases using Leishmania tarentolae and Trypanosoma brucei mitochondrial extracts (11,12). The activity sedimented around 20 -25S and migrated as a single band in a native gel (13). A ϳ20S multiprotein complex that showed both U-insertion and U-deletion in vitro activities was also purified from T. brucei mitochondria by column chromatography. An improved isolation of the L-complex from L. tarentolae was achieved by the tandem affinity purification procedure (TA P) (14); TA P-tagged plasmidexpressed REL1 became incorporated into the L-complex in vivo, allowing isolation of tagged complexes.Fourteen proteins were initially identified by mass spectrometry of the L. tarentolae REL1-TAP pull-down (14). All of the L. tarentolae proteins had homologs in T. brucei, and several additional proteins found in the T. brucei L-complex (15) were later identified in L. tarentolae (Tables S1 and S2).Two 3-prot...

show abstract

mentioning

confidence: 99%

Structure of the core editing complex (L-complex) involved in uridine insertion/deletion RNA editing in trypanosomatid mitochondria

Li¹,

Hui

et al. 2009

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

View full text Add to dashboard Cite

show abstract

“…Gene expression in the kinetoplast-mitochondrion of trypanosomatid protists involves a unique post-transcriptional mRNA maturation process, termed RNA editing, whereby Uresidues are inserted to or deleted from a pre-edited transcript, and a translatable reading frame is thus created (1)(2)(3)(4)(5). 12 of the 18 protein-coding genes in the maxicircle component of the mitochondrial or kinetoplast DNA in Trypanosoma brucei or Leishmania tarentolae encode transcripts that require varying degrees of editing for translation competence (6).…”

mentioning

confidence: 99%

The Effect of RNA Interference Down-regulation of RNA Editing 3′-Terminal Uridylyl Transferase (TUTase) 1 on Mitochondrial de Novo Protein Synthesis and Stability of Respiratory Complexes in Trypanosoma brucei

Neboháčová

Maslov

Falick

et al. 2004

Journal of Biological Chemistry

Self Cite

View full text Add to dashboard Cite

Inhibition of RNA editing by down-regulation of expression of the mitochondrial RNA editing TUTase 1 by RNA interference had profound effects on kinetoplast biogenesis in Trypanosoma brucei procyclic cells. De novo synthesis of the apocytochrome b and cytochrome oxidase subunit I proteins was no longer detectable after 3 days of RNAi. The effect on protein synthesis correlated with a decline in the levels of the assembled mitochondrial respiratory complexes III and IV, and also cyanide-sensitive oxygen uptake. The steady-state levels of nuclear-encoded subunits of complexes III and IV were also significantly decreased. Because the levels of the corresponding mRNAs were not affected, the observed effect was likely due to an increased turnover of these imported mitochondrial proteins. This induced protein degradation was selective for components of complexes III and IV, because little effect was observed on components of the F 1 ⅐F 0 -ATPase complex and on several other mitochondrial proteins.Gene expression in the kinetoplast-mitochondrion of trypanosomatid protists involves a unique post-transcriptional mRNA maturation process, termed RNA editing, whereby Uresidues are inserted to or deleted from a pre-edited transcript, and a translatable reading frame is thus created (1-5). 12 of the 18 protein-coding genes in the maxicircle component of the mitochondrial or kinetoplast DNA in Trypanosoma brucei or Leishmania tarentolae encode transcripts that require varying degrees of editing for translation competence (6). These include mRNAs for apocytochrome b (Cyb), 1 subunits II and III of cytochrome c oxidase (COII and COIII, respectively), subunit 6 of ATPase (6), several subunits of NADH dehydrogenase, and a few others. Some transcripts, such as the cytochrome oxidase subunit I (COI) mRNA, do not require editing for translation. The mechanism of editing includes cleavage of RNA at specific editing sites, and the addition or deletion of a defined number of U-residues followed by religation (7,8). The reactions are performed by large protein complexes which include a 3Ј terminal uridylyl transferase (TUTase), an RNA ligase, endo-and exonuclease activities, as well as several additional auxiliary factors of undefined function (9 -12). Subcomplexes exist that contain subsets of proteins and which may specialize in Uaddition or U-deletion (13). The information for selection of editing sites resides in small "guide" RNAs that are mainly encoded in the kinetoplast DNA minicircles (7, 14). These RNA molecules have 3Ј oligo-U tails (15), which are added posttranscriptionally by the RET1 TUTase (16, 17). Down-regulation of RET1 expression by RNA interference (RNAi) results in a decrease in the steady-state abundance of edited transcripts (16) because of an effect on the 3Ј oligo-U tail of the guide RNAs (37).Although editing provides translatable transcripts, inhibition of editing should affect protein synthesis and, consequently, the assembly of respiratory complexes in the kinetoplast. The de novo synthesis of COI and Cyb po...

show abstract

“…These proteins differ in size and quaternary structure. The RET1 from Leishmania tarentolae is a tetramer of 121-kDa subunits, and the RET2 is a 57-kDa integral component of the multiprotein core-editing L-complex (18,19). Although both TUTases utilize UTP as the preferred nucleotide, RET1 adds hundreds of uridines to an RNA primer in vitro, whereas RET2 transfers mainly one residue.…”

mentioning

confidence: 99%

RNA-editing Terminal Uridylyl Transferase 1

Aphasizheva

Aphasizhev

Simpson

2004

Journal of Biological Chemistry

Self Cite

View full text Add to dashboard Cite

The catalytic, RNA-binding and oligomerization domains of the RNA-editing terminal uridylyl transferase 1 (RET1) from Leishmania tarentolae mitochondria were characterized by mutational analysis. Significant N-and C-terminal portions of the protein were found to be dispensable for UTP polymerization in vitro. Changes of conserved amino acids in the active site demonstrated a general similarity of sugar-phosphate moiety recognition of the incoming ribonucleotide triphosphate by RET1 and eukaryotic poly(A) polymerases. Overlapping RNA-binding and oligomerization regions were mapped to the C-terminal region, which is conserved only among trypanosomatid RET1 enzymes. In the absence of an RNA primer, RET1 can use UTP itself to initiate nucleotide transfer and produce poly(U) molecules of several hundred nucleotides. An N-terminal zinc finger motif is essential for enzyme activity; deletion of this motif or chelation of zinc inhibits activity.Transfer of a nucleotide to an acceptor hydroxyl group is a fundamental chemical reaction involved in a variety of biological processes. Among many enzyme superfamilies that perform this reaction, the DNA polymerase ␤-type group of nucleotidyltransferases is one of most widespread. Members of this group are involved in such diverse pathways as DNA repair, RNA processing, antibiotic resistance, and signal transduction (1). They include poly(A) polymerase (PAP), 1 CCA-adding enzymes, terminal deoxynucleotidyl transferase, kanamycin nucleotidyltransferase, and others. A characteristic feature of the superfamily is the presence of conserved catalytic domain with three metal-coordinating invariant carboxylates and a helical turn motif hG(G/S) 9 -13 XDh(D/E)h (where X represents any amino acid, and h is a hydrophobic amino acid) (2). Two classifications have been proposed based on sequence similarities. The class I subfamily includes the archaeal CCA-adding enzyme, eukaryotic PAP, DNA polymerase ␤, terminal deoxynucleotidyltransferase, and kanamycin nucleotidyltransferase, whereas the class II subfamily contains eubacterial and eukaryotic CCA-adding enzymes and eubacterial PAP (3). Aravind and Koonin (1) expanded the DNA polymerase ␤-type nucleotidyltransferase superfamily to contain several more groups of proteins and divided the enlarged set into nine families.Until recently, PAPs were the only known template-independent enzymes that exhibited specificity toward a particular nucleotide in an otherwise typical polymerization reaction. Atomic structures have been solved for yeast (4) and bovine (5) PAP, and several conserved amino acids were proposed to direct the specificity of the enzyme for ATP, albeit not by direct contacts between the nucleotide base and active center residues.Terminal uridylyltransferases (TUTases), enzymatic activities that add UMP residues to the 3Ј-hydroxyl group of RNA, have been described in mammalian cells (6 -8), plants (9), and trypanosomes (10, 11). No such activity has yet been found in prokaryotes. A biochemical analysis of protein complexes involved i...

show abstract

Uridine insertion/deletion RNA editing in trypanosome mitochondria: A complex business

Cited by 154 publications

References 74 publications

Structure of the core editing complex (L-complex) involved in uridine insertion/deletion RNA editing in trypanosomatid mitochondria

Structure of the core editing complex (L-complex) involved in uridine insertion/deletion RNA editing in trypanosomatid mitochondria

The Effect of RNA Interference Down-regulation of RNA Editing 3′-Terminal Uridylyl Transferase (TUTase) 1 on Mitochondrial de Novo Protein Synthesis and Stability of Respiratory Complexes in Trypanosoma brucei

RNA-editing Terminal Uridylyl Transferase 1

Contact Info

Product

Resources

About