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Alternative mRNA editing in trypanosomes is extensive and may contribute to mitochondrial protein diversity.

Ochsenreiter T, Cipriano M, Hajduk SL - PLoS ONE (2008)

Bottom Line: To extend the analysis of alternative editing in Trypanosoma brucei we have constructed libraries with over 1100 full-length mitochondrial cDNAs and the sequences of over 1200 gRNA genes.Several gRNAs potentially responsible for the alternative editing of these mRNAs were also identified.These findings show that alternative editing of mitochondrial mRNAs is common in T. brucei and expands the diversity of mitochondrial proteins in these organisms.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biology, University of Georgia, Athens, Georgia, USA.

ABSTRACT
The editing of trypanosome mitochondrial mRNAs produces transcripts necessary for mitochondrial functions including electron transport and oxidative phosphorylation. Precursor-mRNAs are often extensively edited by specific uridine insertion or deletion that is directed by small guide RNAs (gRNAs). Recently, it has been shown that cytochrome c oxidase subunit III (COXIII) mRNAs can be alternatively edited to encode a novel mitochondrial membrane protein composed of a unique hydrophilic N-terminal sequence of unknown function and the C-terminal hydrophobic segment of COXIII. To extend the analysis of alternative editing in Trypanosoma brucei we have constructed libraries with over 1100 full-length mitochondrial cDNAs and the sequences of over 1200 gRNA genes. Using this data, we show that alternative editing of COXIII, ATPase subunit 6 (A6), and NADH dehydrogenase subunits 7, 8 and 9 (ND7, 8, 9) mRNAs can produce novel open reading frames (ORFs). Several gRNAs potentially responsible for the alternative editing of these mRNAs were also identified. These findings show that alternative editing of mitochondrial mRNAs is common in T. brucei and expands the diversity of mitochondrial proteins in these organisms.

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Alternative editing of A6 mRNA.A) Alignment of the predicted amino acid sequence of A6 and the alternatively edited cDNA A6-D08. B) Alignment of the RNA sequences for A6, and A6-D08 from the alternatively edited regions I, and II are shown below the bar. In black are the pre-edited residues, in grey the bona fide edited residues and in red are the alternatively edited residues from regions I–III.Bar represents the mRNA sequence of A6-D08. The UUG initiation codon is shown in black. Alignment of the RNA sequence of A6 and A6-D08 from the alternatively edited regions I, II and III (red). Depicted above the RNA sequences are the corresponding amino acid sequences of D08 and A6. The predicted gRNA shows perfect complementarity (allowing for G:U) to A6-D08 over 40 base pairs while having two mismatches to the A6 sequence at position 26-27 of the gRNA. Vertical bars indicate A:U or G:C base pairing; crosses indicates G:U base pairing. Red Us indicate alternatively inserted Us when compared with the A6 sequence. Star shows termination codon in the amino acid sequence.
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pone-0001566-g005: Alternative editing of A6 mRNA.A) Alignment of the predicted amino acid sequence of A6 and the alternatively edited cDNA A6-D08. B) Alignment of the RNA sequences for A6, and A6-D08 from the alternatively edited regions I, and II are shown below the bar. In black are the pre-edited residues, in grey the bona fide edited residues and in red are the alternatively edited residues from regions I–III.Bar represents the mRNA sequence of A6-D08. The UUG initiation codon is shown in black. Alignment of the RNA sequence of A6 and A6-D08 from the alternatively edited regions I, II and III (red). Depicted above the RNA sequences are the corresponding amino acid sequences of D08 and A6. The predicted gRNA shows perfect complementarity (allowing for G:U) to A6-D08 over 40 base pairs while having two mismatches to the A6 sequence at position 26-27 of the gRNA. Vertical bars indicate A:U or G:C base pairing; crosses indicates G:U base pairing. Red Us indicate alternatively inserted Us when compared with the A6 sequence. Star shows termination codon in the amino acid sequence.

Mentions: Analysis of 69 full length A6 cDNAs also identified three alternatively edited cDNAs of identical sequence, designated A6-D08. This sequence contained an ORF that differed from the A6 consensus sequence (Figure 5A). The ORF was initiated by an alternative UUG start codon at nucleotide position 40 and terminated by the bona fide A6 stop codon. The mRNA sequence of A6-D08 was identical to the consensus for A6 at nucleotides 323 to 517. The 5′ 208 nucleotides were the pre-edited sequence encoded by the A6 gene while three regions (I, 260–265; II, 323 and III 517–535 Table 1) were edited to a unique sequence. The formation of the A6-D08 ORF requires the correct alignment of three distinct ORFs encoded by pre-edited, alternatively edited and fully edited A6 at two junctions, one between the pre-edited and edited sequence and the second at the alternatively edited region II (Figure 5B). When the corresponding amino acid sequence was compared to the public databases no sequences with significant similarity were found. We have identified a gRNA gene to the alternatively edited region I of A6-D08 mRNA. The gRNA is predicted to have perfect complementarity over 40 bp to the alternative A6-D08 mRNA from nucleotide position 248-288.


Alternative mRNA editing in trypanosomes is extensive and may contribute to mitochondrial protein diversity.

Ochsenreiter T, Cipriano M, Hajduk SL - PLoS ONE (2008)

Alternative editing of A6 mRNA.A) Alignment of the predicted amino acid sequence of A6 and the alternatively edited cDNA A6-D08. B) Alignment of the RNA sequences for A6, and A6-D08 from the alternatively edited regions I, and II are shown below the bar. In black are the pre-edited residues, in grey the bona fide edited residues and in red are the alternatively edited residues from regions I–III.Bar represents the mRNA sequence of A6-D08. The UUG initiation codon is shown in black. Alignment of the RNA sequence of A6 and A6-D08 from the alternatively edited regions I, II and III (red). Depicted above the RNA sequences are the corresponding amino acid sequences of D08 and A6. The predicted gRNA shows perfect complementarity (allowing for G:U) to A6-D08 over 40 base pairs while having two mismatches to the A6 sequence at position 26-27 of the gRNA. Vertical bars indicate A:U or G:C base pairing; crosses indicates G:U base pairing. Red Us indicate alternatively inserted Us when compared with the A6 sequence. Star shows termination codon in the amino acid sequence.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2215773&req=5

pone-0001566-g005: Alternative editing of A6 mRNA.A) Alignment of the predicted amino acid sequence of A6 and the alternatively edited cDNA A6-D08. B) Alignment of the RNA sequences for A6, and A6-D08 from the alternatively edited regions I, and II are shown below the bar. In black are the pre-edited residues, in grey the bona fide edited residues and in red are the alternatively edited residues from regions I–III.Bar represents the mRNA sequence of A6-D08. The UUG initiation codon is shown in black. Alignment of the RNA sequence of A6 and A6-D08 from the alternatively edited regions I, II and III (red). Depicted above the RNA sequences are the corresponding amino acid sequences of D08 and A6. The predicted gRNA shows perfect complementarity (allowing for G:U) to A6-D08 over 40 base pairs while having two mismatches to the A6 sequence at position 26-27 of the gRNA. Vertical bars indicate A:U or G:C base pairing; crosses indicates G:U base pairing. Red Us indicate alternatively inserted Us when compared with the A6 sequence. Star shows termination codon in the amino acid sequence.
Mentions: Analysis of 69 full length A6 cDNAs also identified three alternatively edited cDNAs of identical sequence, designated A6-D08. This sequence contained an ORF that differed from the A6 consensus sequence (Figure 5A). The ORF was initiated by an alternative UUG start codon at nucleotide position 40 and terminated by the bona fide A6 stop codon. The mRNA sequence of A6-D08 was identical to the consensus for A6 at nucleotides 323 to 517. The 5′ 208 nucleotides were the pre-edited sequence encoded by the A6 gene while three regions (I, 260–265; II, 323 and III 517–535 Table 1) were edited to a unique sequence. The formation of the A6-D08 ORF requires the correct alignment of three distinct ORFs encoded by pre-edited, alternatively edited and fully edited A6 at two junctions, one between the pre-edited and edited sequence and the second at the alternatively edited region II (Figure 5B). When the corresponding amino acid sequence was compared to the public databases no sequences with significant similarity were found. We have identified a gRNA gene to the alternatively edited region I of A6-D08 mRNA. The gRNA is predicted to have perfect complementarity over 40 bp to the alternative A6-D08 mRNA from nucleotide position 248-288.

Bottom Line: To extend the analysis of alternative editing in Trypanosoma brucei we have constructed libraries with over 1100 full-length mitochondrial cDNAs and the sequences of over 1200 gRNA genes.Several gRNAs potentially responsible for the alternative editing of these mRNAs were also identified.These findings show that alternative editing of mitochondrial mRNAs is common in T. brucei and expands the diversity of mitochondrial proteins in these organisms.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biology, University of Georgia, Athens, Georgia, USA.

ABSTRACT
The editing of trypanosome mitochondrial mRNAs produces transcripts necessary for mitochondrial functions including electron transport and oxidative phosphorylation. Precursor-mRNAs are often extensively edited by specific uridine insertion or deletion that is directed by small guide RNAs (gRNAs). Recently, it has been shown that cytochrome c oxidase subunit III (COXIII) mRNAs can be alternatively edited to encode a novel mitochondrial membrane protein composed of a unique hydrophilic N-terminal sequence of unknown function and the C-terminal hydrophobic segment of COXIII. To extend the analysis of alternative editing in Trypanosoma brucei we have constructed libraries with over 1100 full-length mitochondrial cDNAs and the sequences of over 1200 gRNA genes. Using this data, we show that alternative editing of COXIII, ATPase subunit 6 (A6), and NADH dehydrogenase subunits 7, 8 and 9 (ND7, 8, 9) mRNAs can produce novel open reading frames (ORFs). Several gRNAs potentially responsible for the alternative editing of these mRNAs were also identified. These findings show that alternative editing of mitochondrial mRNAs is common in T. brucei and expands the diversity of mitochondrial proteins in these organisms.

Show MeSH
Related in: MedlinePlus