The MRB1 complex functions in kinetoplastid RNA processing

Acestor, Nathalie; Panigrahi, Aswini K.; Carnes, Jason; Zı́ková, Alena; Stuart, Kenneth

doi:10.1261/rna.1353209

Cited by 51 publications

(105 citation statements)

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“…Changes in unedited mRNAs were not statistically significant, as in other studies showing sublethal editing effects, including with our REH2 construct above, where unedited RNAs are not always evidently affected (Supplemental Fig. S4; Hashimi et al 2008;Acestor et al 2009). The never-edited mRNA COI was not affected, while the A6 mRNA may only be slightly decreased.…”

Section: Resultssupporting

confidence: 50%

“…6B,C). Previous studies have used qPCR to detect low-level unprocessed pre-mRNA fragments (Acestor et al 2009). We established similar assays for pre-gRNAs (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…This protein is thought to play a role in the control of gRNA utilization by (or possibly entry into) RECC through undefined mechanisms that may involve annealing and unwinding (Ammerman et al 2010). Another study proposed that its repression leads to accumulation of unprocessed mRNA precursors (Acestor et al 2009), implying that this protein may be multifunctional. Other subunits of MRB complexes have been linked with the binding and stability of gRNAs and the processing of some mRNA and rRNA precursors (Hashimi et al 2008;Panigrahi et al 2008;Weng et al 2008;Acestor et al 2009;Hernandez et al 2010).…”

Section: Discussionmentioning

confidence: 99%

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Guide RNA biogenesis involves a novel RNase III family endoribonuclease inTrypanosoma brucei

Madina¹,

Gokulan²,

Vashisht³

et al. 2011

RNA

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The mitochondrial genome of kinetoplastids, including species of Trypanosoma and Leishmania, is an unprecedented DNA structure of catenated maxicircles and minicircles. Maxicircles represent the typical mitochondrial genome encoding components of the respiratory complexes and ribosomes. However, most mRNA sequences are cryptic, and their maturation requires a unique U insertion/deletion RNA editing. Minicircles encode hundreds of small guide RNAs (gRNAs) that partially anneal with unedited mRNAs and direct the extensive editing. Trypanosoma brucei gRNAs and mRNAs are transcribed as polycistronic precursors, which undergo processing preceding editing; however, the relevant nucleases are unknown. We report the identification and functional characterization of a close homolog of editing endonucleases, mRPN1 (mitochondrial RNA precursor-processing endonuclease 1), which is involved in gRNA biogenesis. Recombinant mRPN1 is a dimeric dsRNAdependent endonuclease that requires Mg 2+ , a critical catalytic carboxylate, and generates 2-nucleotide 39 overhangs. The cleavage specificity of mRPN1 is reminiscent of bacterial RNase III and thus is fundamentally distinct from editing endonucleases, which target a single scissile bond just 59 of short duplexes. An inducible knockdown of mRPN1 in T. brucei results in loss of gRNA and accumulation of precursor transcripts (pre-gRNAs), consistent with a role of mRPN1 in processing. mRPN1 stably associates with three proteins previously identified in relatively large complexes that do not contain mRPN1, and have been linked with multiple aspects of mitochondrial RNA metabolism. One protein, TbRGG2, directly binds mRPN1 and is thought to modulate gRNA utilization by editing complexes. The proposed participation of mRPN1 in processing of polycistronic RNA and its specific protein interactions in gRNA expression are discussed.

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Section: Resultssupporting

confidence: 50%

“…6B,C). Previous studies have used qPCR to detect low-level unprocessed pre-mRNA fragments (Acestor et al 2009). We established similar assays for pre-gRNAs (Fig.…”

Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

See 1 more Smart Citation

Guide RNA biogenesis involves a novel RNase III family endoribonuclease inTrypanosoma brucei

Madina¹,

Gokulan²,

Vashisht³

et al. 2011

RNA

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show abstract

“…The editing reaction is mediated by a holoenzyme or "editosome" (4). The core component of the editosome is the RECC (RNA editing core complex) (5) that contains approximately [15][16][17][18][19][20] polypeptides (2,(6)(7)(8)(9). Eight RECC proteins have been so far functionally characterized.…”

mentioning

confidence: 99%

“…These include the MRP1/2 complex (11, 12) and the guide RNA binding complex (GRBC)/mitochondrial RNA binding (MRB1) complex (13)(14)(15)(16). Heterogeneous-sized high molecular weight complexes (L* or RECC*) consisting of the RECC together with several RNA-linked complexes have been visualized by electrophoresis in blue native gels (4).…”

mentioning

confidence: 99%

Trypanosome REH1 is an RNA helicase involved with the 3′–5′ polarity of multiple gRNA-guided uridine insertion/deletion RNA editing

Herrera

Zhou

et al. 2011

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

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Uridine insertion/deletion RNA editing in kinetoplastid mitochondria corrects encoded frameshifts in mRNAs. The genetic information for editing resides in small guide RNAs (gRNAs), which form anchor duplexes just downstream of an editing site and mediate editing within a single editing "block." Many mRNAs require multiple gRNAs; the observed overall 3′ to 5′ polarity of editing is determined by the formation of upstream mRNA anchors by downstream editing. Hel61, a mitochondrial DEAD-box protein, was previously shown to be involved in RNA editing, but the functional role was not clear. Here we report that down-regulation of Hel61 [renamed REH1 (RNA editing helicase 1)] expression in Trypanosoma brucei selectively affects editing mediated by two or more overlapping gRNAs but has no effect on editing within a single block. Down-regulation produces an increased abundance of the gRNA/edited mRNA duplex for the first editing block of the A6 mRNA. Recombinant REH1 has an ATP-dependent double strand RNA unwinding activity in vitro with a model gRNA-mRNA duplex. These data indicate that REH1 is involved in gRNA displacement either directly by unwinding the gRNA/edited mRNA duplex or indirectly, to allow the 5′ adjacent upstream gRNA to form an anchor duplex with the edited mRNA to initiate another block of editing. Purified tagged REH1 is associated with the RNA editing core complex by RNA linkers and a colocalization of REH1, REL1, and two kinetoplast ribosomal proteins with the kinetoplast DNA was observed by immunofluorescence, suggesting that editing, transcription, and translation may be functionally linked.Leishmania | RNAi | tandem affinity purification | streptavidin binding and protein A purification

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Trypanosome RNA editing: the complexity of getting U in and taking U out

2015

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RNA editing, which adds sequence information to RNAs posttranscriptionally, is a widespread phenomenon throughout eukaryotes. The most complex form of this process is the uridine (U) insertion/deletion editing that occurs in the mitochondria of kinetoplastid protists. RNA editing in these flagellates is specified by trans acting guide RNAs and entails the insertion of hundreds and deletion of dozens of U residues from mitochondrial RNAs to produce mature, translatable mRNAs. An emerging model indicates that the machinery required for trypanosome RNA editing is much more complicated than previously appreciated. A family of RNA editing core complexes (RECCs), which contain the required enzymes and several structural proteins, catalyze cycles of U insertion and deletion. A second, dynamic multi-protein complex, the Mitochondrial RNA Binding 1 (MRB1) complex, has recently come to light as another essential component of the trypanosome RNA editing machinery. MRB1 likely serves as the platform for kinetoplastid RNA editing, and plays critical roles in RNA utilization and editing processivity. MRB1 also appears to act as a hub for coordination of RNA editing with additional mitochondrial RNA processing events. This review highlights the current knowledge regarding the complex molecular machinery involved in trypanosome RNA editing.

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The MRB1 complex functions in kinetoplastid RNA processing

Cited by 51 publications

References 50 publications

Guide RNA biogenesis involves a novel RNase III family endoribonuclease inTrypanosoma brucei

Guide RNA biogenesis involves a novel RNase III family endoribonuclease inTrypanosoma brucei

Trypanosome REH1 is an RNA helicase involved with the 3′–5′ polarity of multiple gRNA-guided uridine insertion/deletion RNA editing

Trypanosome RNA editing: the complexity of getting U in and taking U out

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