Structure and intrinsic disorder of the proteins of the<i>Trypanosoma brucei</i> editosome

Czerwoniec, Anna; Kasprzak, J.; Bytner, Patrycja; Dobrychłop, Mateusz; Bujnicki, Janusz M.

doi:10.1016/j.febslet.2015.07.026

Cited by 7 publications

(18 citation statements)

References 62 publications

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“…Intralinks within KREPB8 and KREPB10 are consistent with the modeled structures of the newly identified domains (SI Appendix, Fig. S5) and with previous modeling of editosome proteins that also detected RNase III domains in KREPB6 and KREPB7 (56). These degenerate RNase III domains are therefore also candidates for functioning as intermolecular heterodimers with their partner endonucleases, which requires further validation by alternative approaches.…”

supporting

confidence: 86%

“…S5 and S6 and Table S7, and Dataset S3) of editosome proteins. Comparative models for parts of all subunits were built using ModPipe (52), HHPred (53), Modeler (54), and Swiss-model (55), which generally identified templates similar to those previously used for comparative modeling for a number of proteins (Dataset S3) (56). The BS3 cross-linker has a linker arm of 11.4 Å when fully extended and can cross-link two residues whose Cα atoms are up to 30 Å apart (57).…”

mentioning

confidence: 99%

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The Architecture of Trypanosoma brucei editosomes

McDermott

Luo

Carnes

et al. 2016

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

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Uridine insertion and deletion RNA editing generates functional mitochondrial mRNAs in Trypanosoma brucei. Editing is catalyzed by three distinct ∼20S editosomes that have a common set of 12 proteins, but are typified by mutually exclusive RNase III endonucleases with distinct cleavage specificities and unique partner proteins. Previous studies identified a network of protein-protein interactions among a subset of common editosome proteins, but interactions among the endonucleases and their partner proteins, and their interactions with common subunits were not identified. Here, chemical cross-linking and mass spectrometry, comparative structural modeling, and genetic and biochemical analyses were used to define the molecular architecture and subunit organization of purified editosomes. We identified intra-and interprotein cross-links for all editosome subunits that are fully consistent with editosome protein structures and previously identified interactions, which we validated by genetic and biochemical studies. The results were used to create a highly detailed map of editosome protein domain proximities, leading to identification of molecular interactions between subunits, insights into the functions of noncatalytic editosome proteins, and a global understanding of editosome architecture.cross-linking | proteomics | Trypanosoma brucei | RNA editing | editosome T he kinetoplastid Trypanosoma brucei is the causative agent of human African trypanosomiasis, which is transmitted by the tsetse fly and is a health threat to millions of people in subSaharan Africa. Kinetoplastids are named for their distinctive mitochondrial DNA network, known as the kinetoplast DNA, which is composed of interlocked DNA maxicircles and minicircles (1, 2). Several identical maxicircles encode two ribosomal RNAs (rRNAs) and messenger RNAs (mRNAs) for 18 mitochondrial proteins, 12 of which undergo posttranscriptional RNA editing (3-9). RNA editing was first described in kinetoplastids (6,8,9), where it is essential (10) and involves insertion and deletion of uridines (Us) to generate translatable mitochondrial transcripts. The sequence information for editing is specified by ∼60-nucleotide guide RNAs (gRNAs) that are encoded in thousands of heterogeneous minicircles (11)(12)(13)(14)(15)(16)(17). Editing occurs by rounds of coordinated catalytic steps that require a number of different enzymes: cleavage of the mRNA substrate by endonucleases, addition of Us by 3′ terminal uridylytransferase (TUTase) or removal of Us by U-specific 3′ exonuclease (exoUase) at insertion or deletion editing sites, respectively, and rejoining of mRNA fragments by RNA ligases. The enzymes that catalyze RNA editing in T. brucei are in ∼20S editosome complexes that also contain proteins with no known catalytic functions (16,(18)(19)(20)(21)(22)(23)(24)(25)(26)(27)(28).There are three similar versions of ∼20S editosomes that have a common set of 12 proteins (SI Appendix, Fig. S1 and Table S1). However, they are compositionally and functionally distinct in that each...

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supporting

confidence: 86%

mentioning

confidence: 99%

The Architecture of Trypanosoma brucei editosomes

McDermott

Luo

Carnes

et al. 2016

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

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“…1. While the percentage of predicted disorder varies between 25% in TbMP18 and 72% in TbMP81, together, the six proteins are more disordered than the average of all other editosomal proteins 17 . Two of the proteins (TbMP63, TbMP81) even contain more disordered than ordered regions.…”

Section: Introductionmentioning

confidence: 93%

“…As spatial restraints we implemented all above-described OB-fold interaction data as well as all published binary interactions of editosome components including the recent chemical cross-linking data of the T. brucei OB-fold proteins [28][29][30] . Structure coordinates for the individual proteins were taken from Czerwoniec et al, 2015 17 (and references therein) and all disordered regions were simulated as coarse-grained flexible shapes. Three hundred independent simulations (300000 steps each) were performed and the resulting 300 models were clustered as outlined in the Supplemental Experimental Procedures.…”

Section: Modeling the Rna-refolding Domain(s) Of The 20s Editosomementioning

confidence: 99%

“…TbMP stands for Trypanosoma brucei mitochondrial protein followed by a number indicating the calculated molecular mass in kDa. TbMP81, TbMP63 and TbMP42 additionally contain two C2H2-type Zn-fingers or C2H2-Zn-finger-like domains and all six proteins have recently been predicted to harbor long stretches of intrinsically disordered regions (IDR) 17 . The domain structure of the different proteins, a 3Dconsensus model of the OB-fold and a plot of their disorder propensities 18 is summarized in Fig.…”

Section: Introductionmentioning

confidence: 99%

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Surface-driven RNA-refolding by the OB-fold proteins of theTrypanosoma bruceieditosome

Voigt

2017

Preprint

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SUMMARYRNA editing in African trypanosomes represents an RNA-processing reaction that generates functional mitochondrial transcripts from sequence-deficient pre-mRNAs. The reaction is catalyzed by a macromolecular protein complex known as the editosome. Editosomes have been demonstrated to execute RNA-chaperone activity to overcome the highly folded nature of pre-edited substrate mRNAs. The molecular basis of this activity is unknown. Here we test five OB-fold proteins of the editosome as potential candidates. We show that the different proteins interact by hetero-oligomerization and we demonstrate that all proteins execute RNA-chaperone activity. Activity differences correlate with the surface areas of the proteins and map predominantly to the intrinsically disordered subdomains of the polypeptides. To provide a structural context for our findings we present a coarse-grained model of the editosome. The model suggests that an inner core of catalytically active editosome components is separated from an outer shell of IDP-domains that act as RNA-remodeling sites.

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The RNA chaperone activity of the Trypanosoma brucei editosome raises the dynamic of bound pre-mRNAs

Leeder

Voigt

Brecht

et al. 2016

Sci Rep

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Mitochondrial transcript maturation in African trypanosomes requires an RNA editing reaction that is characterized by the insertion and deletion of U-nucleotides into otherwise non-functional mRNAs. The reaction is catalyzed by editosomes and requires guide (g)RNAs as templates. Recent data demonstrate that the binding of pre-edited mRNAs to editosomes is followed by a chaperone-type RNA remodeling reaction. Here we map the changes in RNA folding using selective 2′-hydroxyl acylation analyzed by primer extension (SHAPE). We demonstrate that pre-mRNAs in their free state adopt intricately folded, highly stable 2D-structures. Editosome binding renders the pre-mRNAs to adopt 2D-conformations of reduced stabilities. On average about 30% of the nucleotides in every pre-mRNA are affected with a prevalence for U-nucleotides. The data demonstrate that the chaperone activity acts by increasing the flexibility of U-residues to lower their base-pairing probability. This results in a simplified RNA folding landscape with a reduced energy barrier to facilitate the binding of gRNAs. The data provide a first rational for the enigmatic U-specificity of the editing reaction.

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Structure and intrinsic disorder of the proteins of theTrypanosoma brucei editosome

Cited by 7 publications

References 62 publications

The Architecture of Trypanosoma brucei editosomes

The Architecture of Trypanosoma brucei editosomes

Surface-driven RNA-refolding by the OB-fold proteins of theTrypanosoma bruceieditosome

The RNA chaperone activity of the Trypanosoma brucei editosome raises the dynamic of bound pre-mRNAs

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