A bifunctional tRNA import receptor from
            <i>Leishmania</i>
            mitochondria

Goswami, Srikanta; Dhar, Gunjan; Mukherjee, Saikat; Mahata, Bidesh; Chatterjee, Saikat; Home, Pratik; Adhya, Samit

doi:10.1073/pnas.0510869103

Cited by 51 publications

(91 citation statements)

References 30 publications

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“…One possibility is that BFs require the COII protein either for complex IV function or for an unknown function. Such an unknown function may be analogous to that of ATP synthase complex, since it functions in reverse in BFs compared to PFs (53), and its ␣ subunit has been reported to actively import tRNAs into Leishmania mitochondria (20). Alternatively, the lethality may reflect loss of editing of other mRNAs, as suggested above, or KREN3 may have an additional unknown function such as cleavage of polycistronic maxicircle or minicircle transcripts (21).…”

Section: Discussionmentioning

confidence: 91%

RNA Editing in Trypanosoma brucei Requires Three Different Editosomes

Carnes

Trotter

Peltan

et al. 2008

Molecular and Cellular Biology

106

150

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Trypanosoma brucei has three distinct ϳ20S editosomes that catalyze RNA editing by the insertion and deletion of uridylates. Editosomes with the KREN1 or KREN2 RNase III type endonucleases specifically cleave deletion and insertion editing site substrates, respectively. We report here that editosomes with KREPB2, which also has an RNase III motif, specifically cleave cytochrome oxidase II (COII) pre-mRNA insertion editing site substrates in vitro. Conditional repression and mutation studies also show that KREPB2 is an editing endonuclease specifically required for COII mRNA editing in vivo. Furthermore, KREPB2 expression is essential for the growth and survival of bloodstream forms. Thus, editing in T. brucei requires at least three compositionally and functionally distinct ϳ20S editosomes, two of which distinguish between different insertion editing sites. This unexpected finding reveals an additional level of complexity in the RNA editing process and suggests a mechanism for how the selection of sites for editing in vivo is controlled.RNA editing recodes most of the mitochondrial mRNAs in trypanosomatids by the insertion and deletion of uridine nucleotides (U's) using information provided by guide RNAs (gRNAs) (58). RNA editing is developmentally regulated and is required for parasites that cycle between insect vector and mammalian host (54). Proteins that perform catalytic steps of RNA editing have been identified: endonucleases KREN1 or KREN2 cleave at deletion or insertion sites, respectively; terminal uridylyl transferase (TUTase) KRET2 adds U's at insertion sites; U specific exoribonuclease (exoUase) KREX1 removes U's at deletion sites; and ligases KREL1 or KREL2 rejoin mRNA fragments after U addition or removal (5,13,25,30,34,55,61). These catalytic steps are coordinated by the multiprotein editosomes that sediment at ϳ20S on glycerol gradients (9, 43). KREPB2 was identified as an editosome component by mass spectrometry, and found to contain an RNase III motif harboring amino acids that are critical for catalysis, a U1 Zn 2ϩ finger, and double-stranded RNA binding. It has sequence similarity to KREN1 and KREN2, which also contain these three motifs (23,41,64). These data strongly suggest an RNA editing endonuclease role for KREPB2.KREN1 and KREN2 were shown to be RNA editing endonucleases using both RNA interference (RNAi) and conditional knockout cell lines (5, 61). Both KREN1 and KREN2 are essential for the normal growth of PF and BF cells, and repression of their expression by either method led to dramatic growth defects or death, respectively. Such repression of KREN1 eliminated in vitro cleavage by ϳ20S editosomes of a deletion site in a synthetic substrate modeled on ATPase subunit 6 (A6) pre-mRNA but did not alter cleavage of an insertion site in a substrate derived from the same mRNA. These studies and other data have shown that KREN1 is an editing endonuclease with a preference for sites from which U's are deleted (29). Similarly, repression of KREN2 eliminated in vitro cleavage at insertion ed...

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Section: Discussionmentioning

confidence: 91%

RNA Editing in Trypanosoma brucei Requires Three Different Editosomes

Carnes

Trotter

Peltan

et al. 2008

Molecular and Cellular Biology

106

150

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“…Whether this putative receptor belongs to the TOM complex or not is an interesting question to answer in the future. Furthermore, an RNA import complex, including two bifunctional respiratory proteins, isolated from L. tropica inner mitochondrial membrane is implicated in tRNA import (12,13). However, because of the conflicting data reported so far (see above), identification of import factors in other trypanosomatids might help in understanding the complexity of mitochondrial tRNA import.…”

Section: Discussionmentioning

confidence: 99%

“…A 15-kDa putative import tRNA receptor has been reported in L. tropica, but this protein remains to be identified (10). More recently, it has been shown in L. tropica that a multisubunit RNA import complex (RIC) located on the inner mitochondrial membrane is implicated in tRNA import (11) and two subunits, RIC1 (a structural homologue to the ␣ subunit of F1 ATP synthase) and RIC8 (an homologue to subunit 6b of ubiquinol cytochrome c reductase) were identified (12,13). However, considering the contradictory data obtained so far, a deeper understanding of the import factors involved in different trypanosomatids will be important in the future.…”

mentioning

confidence: 99%

The voltage-dependent anion channel, a major component of the tRNA import machinery in plant mitochondria

Salinas

Duchêne

Delage

et al. 2006

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

111

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In plants, as in most eukaryotic cells, import of nuclear-encoded cytosolic tRNAs is an essential process for mitochondrial biogenesis. Despite its broad occurrence, the mechanisms governing RNA transport into mitochondria are far less understood than protein import. This article demonstrates by Northwestern and gel-shift experiments that the plant mitochondrial voltage-dependent anion channel (VDAC) protein interacts with tRNA in vitro. It shows also that this porin, known to play a key role in metabolite transport, is a major component of the channel involved in the tRNA translocation step through the plant mitochondrial outer membrane, as supported by inhibition of tRNA import into isolated mitochondria by VDAC antibodies and Ruthenium red. However VDAC is not a tRNA receptor on the outer membrane. Rather, two major components from the TOM (translocase of the outer mitochondrial membrane) complex, namely TOM20 and TOM40, are important for tRNA binding at the surface of mitochondria, suggesting that they are also involved in tRNA import. Finally, we show that proteins and tRNAs are translocated into plant mitochondria by different pathways. Together, these findings identify unexpected components of the tRNA import machinery and suggest that the plant tRNA import pathway has evolved by recruiting multifunctional proteins. mitochondrial porin ͉ cytosolic tRNAs ͉ import factor

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“…The potential problem of an imbalanced matrix tRNA pool is solved by the unique phenomenon of allosteric regulation; type I RNAs stimulate the inner membrane transfer of type II RNAs, whereas type II RNAs inhibit transfer of type I (11,12 (11)(12)(13); in vitro evolution experiments suggest that many more tRNA species belong to these categories (11). The "ping-pong" model (14) postulates that the two types of tRNA bind to different receptors; binding of type I tRNA to its receptor induces a conformational change that is transmitted to the type II receptor, opening up its tRNAbinding site.…”

Section: From the Genetic Engineering Laboratory Indian Institute Ofmentioning

confidence: 99%

“…Two tRNA-binding proteins with the properties of type I and type II receptors are associated with this complex (14). We have recently identified the type I receptor and showed that, in vivo, it is required for import of type I as well as type II tRNAs (13).…”

Section: From the Genetic Engineering Laboratory Indian Institute Ofmentioning

confidence: 99%

An RNA-binding Respiratory Component Mediates Import of Type II tRNAs into Leishmania Mitochondria

Chatterjee

Home

Mukherjee

et al. 2006

Journal of Biological Chemistry

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Transport of tRNAs across the inner mitochondrial membrane of the kinetoplastid protozoon Leishmania requires interactions with specific binding proteins (receptors) in a multi-subunit complex. The allosteric model of import regulation proposes cooperative and antagonistic interactions between two or more receptors with binding specificities for distinct tRNA families (types I and II, respectively). To identify the type II receptor, the gene encoding RIC8A, a subunit of the complex, was cloned. The C-terminal region of RIC8A is homologous to subunit 6b of ubiquinol cytochrome c reductase (respiratory complex III), while the N-terminal region has intrinsic affinity for type II, but not for type I, tRNAs. RIC8A is shared by the import complex and complex III, indicating its bi-functionality, but is assembled differently in the two complexes. Knockdown of RIC8A in Leishmania lowered the mitochondrial content of type II tRNAs but raised that of type I tRNAs, with downstream effects on mitochondrial translation and respiration, and cell death. In RIC8A knockdown cells, a subcomplex was formed that interacted with type I tRNA, but the negative regulation by type II tRNA was lost. Mitochondrial extracts from these cells were defective for type II, but not type I, import; import and regulation were restored by purified RIC8A. These results provide evidence for the relevance of allosteric regulation in vivo and indicate that acquisition of new tRNA-binding domains by ancient respiratory components have played a key role in the evolution of mitochondrial tRNA import.

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A bifunctional tRNA import receptor from Leishmania mitochondria

Cited by 51 publications

References 30 publications

RNA Editing in Trypanosoma brucei Requires Three Different Editosomes

RNA Editing in Trypanosoma brucei Requires Three Different Editosomes

The voltage-dependent anion channel, a major component of the tRNA import machinery in plant mitochondria

An RNA-binding Respiratory Component Mediates Import of Type II tRNAs into Leishmania Mitochondria

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