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Complete mitochondrial genome sequence of three Tetrahymena species reveals mutation hot spots and accelerated nonsynonymous substitutions in Ymf genes.

Moradian MM, Beglaryan D, Skozylas JM, Kerikorian V - PLoS ONE (2007)

Bottom Line: We also found distinct features in Mt genome of T.paravorax despite similar genome organization among these approximately 47 kb long linear genomes.Importantly, nucleotide substitution types and rates suggest possible reasons for not being able to find homologues for Ymf genes.Additionally, comparative genomic analysis of complete Mt genomes is essential in identifying biologically significant motifs such as control regions.

View Article: PubMed Central - PubMed

Affiliation: Department of Ecology and Evolutionary Biology, University of California Los Angeles, Los Angeles, California, United States of America. mmoradia@ucla.edu

ABSTRACT
The ciliate Tetrahymena, a model organism, contains divergent mitochondrial (Mt) genome with unusual properties, where half of its 44 genes still remain without a definitive function. These genes could be categorized into two major groups of KPC (known protein coding) and Ymf (genes without an identified function). To gain insights into the mechanisms underlying gene divergence and molecular evolution of Tetrahymena (T.) Mt genomes, we sequenced three Mt genomes of T.paravorax, T.pigmentosa, and T.malaccensis. These genomes were aligned and the analyses were carried out using several programs that calculate distance, nucleotide substitution (dn/ds), and their rate ratios (omega) on individual codon sites and via a sliding window approach. Comparative genomic analysis indicated a conserved putative transcription control sequence, a GC box, in a region where presumably transcription and replication initiate. We also found distinct features in Mt genome of T.paravorax despite similar genome organization among these approximately 47 kb long linear genomes. Another significant finding was the presence of at least one or more highly variable regions in Ymf genes where majority of substitutions were concentrated. These regions were mutation hotspots where elevated distances and the dn/ds ratios were primarily due to an increase in the number of nonsynonymous substitutions, suggesting relaxed selective constraint. However, in a few Ymf genes, accelerated rates of nonsynonymous substitutions may be due to positive selection. Similarly, on protein level the majority of amino acid replacements occurred in these regions. Ymf genes comprise half of the genes in Tetrahymena Mt genomes, so understanding why they have not been assigned definitive functions is an important aspect of molecular evolution. Importantly, nucleotide substitution types and rates suggest possible reasons for not being able to find homologues for Ymf genes. Additionally, comparative genomic analysis of complete Mt genomes is essential in identifying biologically significant motifs such as control regions.

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Gene map of the T.paravorax Mt genome.Arrows denote direction of transcription. Intergenic region between Cob and Ymf77 genes is considered to contain a putative control region. The trnK* represents the pseudo tRNA in T.paravorax Mt genome.
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pone-0000650-g001: Gene map of the T.paravorax Mt genome.Arrows denote direction of transcription. Intergenic region between Cob and Ymf77 genes is considered to contain a putative control region. The trnK* represents the pseudo tRNA in T.paravorax Mt genome.

Mentions: We sequenced the entire Mt genomes of T.paravorax, T.pigmentosa, and T.malaccensis. These Mt genomes, along with the two previously sequenced Mt genomes of T.pyriformis and T. thermophila, are ∼47 kb long, have high A+T content (Table S1), and contain 44 genes. Definitive functions could be assigned to just 22 of them. The remaining 22 were putative proteins with either similar domains to known proteins or no similarity at all. Genome organization and gene arrangements in all five genomes were the same with the exception of an additional putative pseudo-tRNA lysine (K) in T.paravorax, which was located between the 5′ telomere and the inverted repeat region (Figure 1). Pseudo-tRNAs are pseudogenes that cannot form cloverleaf shape structures but can form stem loops (Figure S1). A putative pseudo-lysine-tRNA in T.paravorax Mt genome was located at 5′ end of the genome in the unusually long non-coding sequence (495bp) between the telomere and the large ribosomal subunit. The putative pseudo-lysine-tRNA is located 118 bp away from the telomere. The presence of pseudo-tRNAs is not an unprecedented phenomenon in Mt genomes since pseudo-tRNAs have been reported in Mt genome of plague Thrips imaginis and a few others [19]. In the T.pigmentosa Mt genome the non-coding sequence is absent at the 3′ end of the genome and the non-coding sequence at the 5′ end is only 31 bp. None of the other three species have longer than 65 bp non-coding sequences adjacent to the telomeres. Thus the presence of a long non-coding region with a pseudo-tRNA in T.paravorax may point to the process of elimination and transfer of Mt genes. It is possible that at some point during evolution there was a functional Lysine tRNA adjacent to the telomeres in Mt genome of T.paravorax. Another interesting feature of the T.paravorax Mt genome is a 1 kb sequence in Ymf77 that can be folded into a secondary structure with a very big free energy value, lowest ΔG = −125 Kcal/mole. Such a low ΔG value was not found in homologous region of the other Mt genomes. The A+T content of this T.paravorax region was 96.5% and its secondary structure made it difficult to clone or PCR amplify the region directly from intact Mt DNA. PCR amplification was eventually achieved for a restriction fragment of about 3.9 kb containing this region, after boiling the DNA in 1% SDS.


Complete mitochondrial genome sequence of three Tetrahymena species reveals mutation hot spots and accelerated nonsynonymous substitutions in Ymf genes.

Moradian MM, Beglaryan D, Skozylas JM, Kerikorian V - PLoS ONE (2007)

Gene map of the T.paravorax Mt genome.Arrows denote direction of transcription. Intergenic region between Cob and Ymf77 genes is considered to contain a putative control region. The trnK* represents the pseudo tRNA in T.paravorax Mt genome.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC1919467&req=5

pone-0000650-g001: Gene map of the T.paravorax Mt genome.Arrows denote direction of transcription. Intergenic region between Cob and Ymf77 genes is considered to contain a putative control region. The trnK* represents the pseudo tRNA in T.paravorax Mt genome.
Mentions: We sequenced the entire Mt genomes of T.paravorax, T.pigmentosa, and T.malaccensis. These Mt genomes, along with the two previously sequenced Mt genomes of T.pyriformis and T. thermophila, are ∼47 kb long, have high A+T content (Table S1), and contain 44 genes. Definitive functions could be assigned to just 22 of them. The remaining 22 were putative proteins with either similar domains to known proteins or no similarity at all. Genome organization and gene arrangements in all five genomes were the same with the exception of an additional putative pseudo-tRNA lysine (K) in T.paravorax, which was located between the 5′ telomere and the inverted repeat region (Figure 1). Pseudo-tRNAs are pseudogenes that cannot form cloverleaf shape structures but can form stem loops (Figure S1). A putative pseudo-lysine-tRNA in T.paravorax Mt genome was located at 5′ end of the genome in the unusually long non-coding sequence (495bp) between the telomere and the large ribosomal subunit. The putative pseudo-lysine-tRNA is located 118 bp away from the telomere. The presence of pseudo-tRNAs is not an unprecedented phenomenon in Mt genomes since pseudo-tRNAs have been reported in Mt genome of plague Thrips imaginis and a few others [19]. In the T.pigmentosa Mt genome the non-coding sequence is absent at the 3′ end of the genome and the non-coding sequence at the 5′ end is only 31 bp. None of the other three species have longer than 65 bp non-coding sequences adjacent to the telomeres. Thus the presence of a long non-coding region with a pseudo-tRNA in T.paravorax may point to the process of elimination and transfer of Mt genes. It is possible that at some point during evolution there was a functional Lysine tRNA adjacent to the telomeres in Mt genome of T.paravorax. Another interesting feature of the T.paravorax Mt genome is a 1 kb sequence in Ymf77 that can be folded into a secondary structure with a very big free energy value, lowest ΔG = −125 Kcal/mole. Such a low ΔG value was not found in homologous region of the other Mt genomes. The A+T content of this T.paravorax region was 96.5% and its secondary structure made it difficult to clone or PCR amplify the region directly from intact Mt DNA. PCR amplification was eventually achieved for a restriction fragment of about 3.9 kb containing this region, after boiling the DNA in 1% SDS.

Bottom Line: We also found distinct features in Mt genome of T.paravorax despite similar genome organization among these approximately 47 kb long linear genomes.Importantly, nucleotide substitution types and rates suggest possible reasons for not being able to find homologues for Ymf genes.Additionally, comparative genomic analysis of complete Mt genomes is essential in identifying biologically significant motifs such as control regions.

View Article: PubMed Central - PubMed

Affiliation: Department of Ecology and Evolutionary Biology, University of California Los Angeles, Los Angeles, California, United States of America. mmoradia@ucla.edu

ABSTRACT
The ciliate Tetrahymena, a model organism, contains divergent mitochondrial (Mt) genome with unusual properties, where half of its 44 genes still remain without a definitive function. These genes could be categorized into two major groups of KPC (known protein coding) and Ymf (genes without an identified function). To gain insights into the mechanisms underlying gene divergence and molecular evolution of Tetrahymena (T.) Mt genomes, we sequenced three Mt genomes of T.paravorax, T.pigmentosa, and T.malaccensis. These genomes were aligned and the analyses were carried out using several programs that calculate distance, nucleotide substitution (dn/ds), and their rate ratios (omega) on individual codon sites and via a sliding window approach. Comparative genomic analysis indicated a conserved putative transcription control sequence, a GC box, in a region where presumably transcription and replication initiate. We also found distinct features in Mt genome of T.paravorax despite similar genome organization among these approximately 47 kb long linear genomes. Another significant finding was the presence of at least one or more highly variable regions in Ymf genes where majority of substitutions were concentrated. These regions were mutation hotspots where elevated distances and the dn/ds ratios were primarily due to an increase in the number of nonsynonymous substitutions, suggesting relaxed selective constraint. However, in a few Ymf genes, accelerated rates of nonsynonymous substitutions may be due to positive selection. Similarly, on protein level the majority of amino acid replacements occurred in these regions. Ymf genes comprise half of the genes in Tetrahymena Mt genomes, so understanding why they have not been assigned definitive functions is an important aspect of molecular evolution. Importantly, nucleotide substitution types and rates suggest possible reasons for not being able to find homologues for Ymf genes. Additionally, comparative genomic analysis of complete Mt genomes is essential in identifying biologically significant motifs such as control regions.

Show MeSH
Related in: MedlinePlus