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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 ND7 mRNAs.A) Alignment of the predicted amino acid sequence of ND7 and the alternatively edited ND7 cDNAs ND7-G10 and ND7-N12. Amino acids changed by alternative editing are in blue. B) Bar depicts the alternatively edited mRNA sequences ND7-G10 and ND7-N12. Alignment of the RNA sequences for ND7, ND7-N12 and ND7-G10 from the alternatively edited regions I, II and III 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. Above the RNA sequences are the corresponding amino acid sequences. A predicted gRNA for N12 shows perfect complementarity (allowing for G:U) to ND7-N12 over 30 base pairs while having mismatches to the ND7 sequence at nucleotide positions 10 and 14 of the gRNA. Vertical bars indicate A:U or G:C base pairing; crosses indicates G:U base pairing. Underlined sequence shows UAG termination codon. Star depicts amber codon in the amino acid sequence. Red Us indicate alternatively inserted/not inserted Us when compared with the ND7 sequence.
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pone-0001566-g001: Alternative editing of ND7 mRNAs.A) Alignment of the predicted amino acid sequence of ND7 and the alternatively edited ND7 cDNAs ND7-G10 and ND7-N12. Amino acids changed by alternative editing are in blue. B) Bar depicts the alternatively edited mRNA sequences ND7-G10 and ND7-N12. Alignment of the RNA sequences for ND7, ND7-N12 and ND7-G10 from the alternatively edited regions I, II and III 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. Above the RNA sequences are the corresponding amino acid sequences. A predicted gRNA for N12 shows perfect complementarity (allowing for G:U) to ND7-N12 over 30 base pairs while having mismatches to the ND7 sequence at nucleotide positions 10 and 14 of the gRNA. Vertical bars indicate A:U or G:C base pairing; crosses indicates G:U base pairing. Underlined sequence shows UAG termination codon. Star depicts amber codon in the amino acid sequence. Red Us indicate alternatively inserted/not inserted Us when compared with the ND7 sequence.

Mentions: To determine whether alternative mRNA editing could produce isoforms of the Complex I subunits we carried out a detailed analysis of cDNA sequence data from extensively edited ND7, ND8 and ND9 mRNAs isolated from the bloodstream developmental stage of T. brucei. The sequence of ND7 mRNA was originally deduced based on the consensus sequence of 77 short cDNAs and direct RNA sequencing [6]. The bona fide ND7 mRNA contained 551 uridines added and 86 uridines deleted by RNA editing to form the 1246 nts mRNA. We have extended the characterization of ND7 RNA editing by detailed analysis of 21 full-length cDNAs and examination of the protein coding potential for each transcript. The sequence of ND7 mRNAs revealed two alternatively edited RNAs with unique ORFs (Figure 1). It was previously noted that differential editing of ND7 mRNAs could lead to diverse ORFs and dramatic differences in editing correlated with the developmental stage of the parasite [6]. We have verified similar changes leading to frame shifting and amino acid substitutions in the ND7 mRNAs in bloodstream T. brucei.


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 ND7 mRNAs.A) Alignment of the predicted amino acid sequence of ND7 and the alternatively edited ND7 cDNAs ND7-G10 and ND7-N12. Amino acids changed by alternative editing are in blue. B) Bar depicts the alternatively edited mRNA sequences ND7-G10 and ND7-N12. Alignment of the RNA sequences for ND7, ND7-N12 and ND7-G10 from the alternatively edited regions I, II and III 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. Above the RNA sequences are the corresponding amino acid sequences. A predicted gRNA for N12 shows perfect complementarity (allowing for G:U) to ND7-N12 over 30 base pairs while having mismatches to the ND7 sequence at nucleotide positions 10 and 14 of the gRNA. Vertical bars indicate A:U or G:C base pairing; crosses indicates G:U base pairing. Underlined sequence shows UAG termination codon. Star depicts amber codon in the amino acid sequence. Red Us indicate alternatively inserted/not inserted Us when compared with the ND7 sequence.
© Copyright Policy
Related In: Results  -  Collection

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

pone-0001566-g001: Alternative editing of ND7 mRNAs.A) Alignment of the predicted amino acid sequence of ND7 and the alternatively edited ND7 cDNAs ND7-G10 and ND7-N12. Amino acids changed by alternative editing are in blue. B) Bar depicts the alternatively edited mRNA sequences ND7-G10 and ND7-N12. Alignment of the RNA sequences for ND7, ND7-N12 and ND7-G10 from the alternatively edited regions I, II and III 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. Above the RNA sequences are the corresponding amino acid sequences. A predicted gRNA for N12 shows perfect complementarity (allowing for G:U) to ND7-N12 over 30 base pairs while having mismatches to the ND7 sequence at nucleotide positions 10 and 14 of the gRNA. Vertical bars indicate A:U or G:C base pairing; crosses indicates G:U base pairing. Underlined sequence shows UAG termination codon. Star depicts amber codon in the amino acid sequence. Red Us indicate alternatively inserted/not inserted Us when compared with the ND7 sequence.
Mentions: To determine whether alternative mRNA editing could produce isoforms of the Complex I subunits we carried out a detailed analysis of cDNA sequence data from extensively edited ND7, ND8 and ND9 mRNAs isolated from the bloodstream developmental stage of T. brucei. The sequence of ND7 mRNA was originally deduced based on the consensus sequence of 77 short cDNAs and direct RNA sequencing [6]. The bona fide ND7 mRNA contained 551 uridines added and 86 uridines deleted by RNA editing to form the 1246 nts mRNA. We have extended the characterization of ND7 RNA editing by detailed analysis of 21 full-length cDNAs and examination of the protein coding potential for each transcript. The sequence of ND7 mRNAs revealed two alternatively edited RNAs with unique ORFs (Figure 1). It was previously noted that differential editing of ND7 mRNAs could lead to diverse ORFs and dramatic differences in editing correlated with the developmental stage of the parasite [6]. We have verified similar changes leading to frame shifting and amino acid substitutions in the ND7 mRNAs in bloodstream T. brucei.

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