Developmental aspects of uridine addition within mitochondrial transcripts of Trypanosoma brucei.

Feagin, Jean E.; Stuart, Kenneth

doi:10.1128/mcb.8.3.1259

Cited by 101 publications

(49 citation statements)

References 25 publications

(49 reference statements)

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“…A number of transcripts are differentially edited between BF and PF cells (3)(4)(5)(6)(7)(72)(73)(74). The molecular basis for this developmental regulation remains unknown, although a growing body of evidence shows that elimination or mutation of the same editosome protein differentially affects RNA editing in BF and PF parasites, and similar results have been observed with other mitochondrial RNA processing protein complexes (36,60,75).…”

Section: Discussionsupporting

confidence: 56%

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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Section: Discussionsupporting

confidence: 56%

The Architecture of Trypanosoma brucei editosomes

McDermott

Luo

Carnes

et al. 2016

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

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“…To contrast A6, 5' CYbUT, which encompasses 88 nt of the 5' end of pre-edited Cytochrome b, and its gRNA, gCYb-558, was the other substrate pair examined. Editing in this mRNA is developmentally regulated with edited message produced only in the procyclic (insect) and stumpy bloodstream stages of the trypanosome life cycle [22,23]. The short (19 nt) editing domain is also purine-rich (95%) however, solution structure probing of this substrate indicates that it forms a very stable stem loop with the anchor sequence hidden within the stem ( Figure 1C) [11].…”

Section: Analysis Of Selected Grna/mrna Pairsmentioning

confidence: 99%

Evidence for U-tail Stabilization of gRNA/mRNA Interactions in Kinetoplastid RNA Editing

Koslowsky¹,

Reifur²,

Yu³

et al. 2004

RNA Biology

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The most dramatic example of RNA editing is found in the mitochondria of trypanosomes. In these organisms, U-insertions/deletions can create mRNAs that are twice as large as the gene that encodes them. Guide RNAs (gRNAs) that are complementary to short stretches of the mature message direct the precise placements of the U residues. The binding of gRNA to mRNA is a fundamental step in RNA editing and understanding the relative importance of the elements that confer affinity and specificity on this interaction is critical to our understanding of the editing process. In this study, we have analyzed the relative binding affinities of two different gRNA/mRNA pairs. The affinity of gA6-14 for its message (ATPase 6) is high, with an apparent K D in the 5-10 nM range. In contrast, gCYb-558 has a low affinity for its cognate mRNA. Deletion of the gRNA U-tail caused a significant reduction in the binding affinity for only the gCYb-558 pair, and was observed only under physiological magnesium conditions. These results indicate that the U-tail contribution can differ substantially between the different gRNA/mRNA pairs. In addition, our results suggest that the efficiency of gRNA/mRNA interaction is highly dependent on thermodynamic parameters determined by the local sequences and their adopted structures surrounding the anchor-binding site.

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“…T. brucei has recently been discovered to possess a unique RNA modification activity which allows internal additions of uridine to mitochondrial transcripts (4,(14)(15)(16). This activity FIG.…”

Section: Resultsmentioning

confidence: 99%

Posttranscriptional regulation of cytochrome c expression during the developmental cycle of Trypanosoma brucei.

Torri¹,

Hajduk²

1988

Mol. Cell. Biol.

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We examined the expression of a nucleus-encoded mitochondrial protein, cytochrome c, during the life cycle of Trypanosoma brucei. The bloodstream forms of T. brucei, the long slender and short stumpy trypanosomes, have inactive mitochondria with no detectable cytochrome-mediated respiration. The insect form of T. brucei, the procyclic trypanosomes, has fully functional mitochondria. Cytochrome c is spectrally undetectable in the bloodstream forms of trypanosomes, but during differentiation to the procyclic form, spectrally detected holo-cytochrome c accumulates rapidly. We have purified T. brucei cytochrome c and raised antibodies that react to both holo- and apo-cytochrome c. In addition, we isolated a partial cDNA to trypanosome cytochrome c. An examination of protein expression and steady-state mRNA levels in T. brucei indicated that bloodstream trypanosomes did not express cytochrome c but maintained significant steady-state levels of cytochrome c mRNA. The results suggest that in T. brucei, cytochrome c is developmentally regulated by a posttranscriptional mechanism which prevents either translation or accumulation of cytochrome c in the bloodstream trypanosomes.

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Developmental aspects of uridine addition within mitochondrial transcripts of Trypanosoma brucei.

Cited by 101 publications

References 25 publications

The Architecture of Trypanosoma brucei editosomes

The Architecture of Trypanosoma brucei editosomes

Evidence for U-tail Stabilization of gRNA/mRNA Interactions in Kinetoplastid RNA Editing

Posttranscriptional regulation of cytochrome c expression during the developmental cycle of Trypanosoma brucei.

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