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Alternative translation initiation codons for the plastid maturase MatK: unraveling the pseudogene misconception in the Orchidaceae.

Barthet MM, Moukarzel K, Smith KN, Patel J, Hilu KW - BMC Evol. Biol. (2015)

Bottom Line: We confirm that MatK protein is expressed and functions in sample orchids currently described as having a matK pseudogene using immunodetection and reverse-transcription methods.We demonstrate using phylogenetic analysis that this alternative initiation codon emerged de novo within the Orchidaceae, with several reversal events at the basal lineage and deep in orchid history.These findings suggest a novel evolutionary shift for expression of matK in the Orchidaceae and support the function of MatK as a group II intron maturase in the plastid genome of land plants including the orchids.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, Coastal Carolina University, Conway, SC, 29526, USA. mbarthet@coastal.edu.

ABSTRACT

Background: The plastid maturase MatK has been implicated as a possible model for the evolutionary "missing link" between prokaryotic and eukaryotic splicing machinery. This evolutionary implication has sparked investigations concerning the function of this unusual maturase. Intron targets of MatK activity suggest that this is an essential enzyme for plastid function. The matK gene, however, is described as a pseudogene in many photosynthetic orchid species due to presence of premature stop codons in translations, and its high rate of nucleotide and amino acid substitution.

Results: Sequence analysis of the matK gene from orchids identified an out-of-frame alternative AUG initiation codon upstream from the consensus initiation codon used for translation in other angiosperms. We demonstrate translation from the alternative initiation codon generates a conserved MatK reading frame. We confirm that MatK protein is expressed and functions in sample orchids currently described as having a matK pseudogene using immunodetection and reverse-transcription methods. We demonstrate using phylogenetic analysis that this alternative initiation codon emerged de novo within the Orchidaceae, with several reversal events at the basal lineage and deep in orchid history.

Conclusion: These findings suggest a novel evolutionary shift for expression of matK in the Orchidaceae and support the function of MatK as a group II intron maturase in the plastid genome of land plants including the orchids.

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Sample alignments showing the consensus initiation codon (cic) and subsequent translation products for MatK in orchids. (a) Nucleotide alignment of various orchid species and the non-orchid monocot species Asparagus aethiopicus, Hordeum vulgare, Saccharum officinarum and Oryza sativa. Asterisks next to species name indicate those noted in GenBank to contain matK as a pseudogene. Black arrow indicates cic used by other monocots and angiosperms for MatK expression. (b) Translated MatK amino acid sequence using the cic. Asterisk in translated amino acid sequence indicates stop codons in the MatK reading frame. Gray shadowing indicates highly conserved sequence among all taxa in the alignment, lighter gray indicates less conserved sequence, white background indicates a lack of conserved sequence. Note: gaps in nucleotide and amino acid alignment differ in relative position due to indel in Palmorchis trilobulata
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Fig1: Sample alignments showing the consensus initiation codon (cic) and subsequent translation products for MatK in orchids. (a) Nucleotide alignment of various orchid species and the non-orchid monocot species Asparagus aethiopicus, Hordeum vulgare, Saccharum officinarum and Oryza sativa. Asterisks next to species name indicate those noted in GenBank to contain matK as a pseudogene. Black arrow indicates cic used by other monocots and angiosperms for MatK expression. (b) Translated MatK amino acid sequence using the cic. Asterisk in translated amino acid sequence indicates stop codons in the MatK reading frame. Gray shadowing indicates highly conserved sequence among all taxa in the alignment, lighter gray indicates less conserved sequence, white background indicates a lack of conserved sequence. Note: gaps in nucleotide and amino acid alignment differ in relative position due to indel in Palmorchis trilobulata

Mentions: Two data sets were generated, one for the molecular and the other for the informatics aspects of this study. In the former case, GenBank nucleotide sequences were obtained for 13 orchid and four related monocot species representing cases where matK was considered as either functional or a pseudogene (Table 1). Translation of matK sequence from the initiation codon used for other monocots (referred to hereafter as the consensus initiation codon, cic) produced full length amino acid sequence for MatK protein from the monocot species Asparagus aethiopicus , Hordeum vulgare, Saccharum officinarum, and Oryza sativa , and three orchid species (Fig. 1). Amino acid sequence from the remaining ten orchid species when translated from the cic displayed premature stop codons suggestive of truncated, non-functional protein (Fig. 1b). Two additional initiation codons were identified upstream of the cic. The first one located −6 bp upstream and in-frame with the cic, was found only in six of the thirteen orchid taxa examined in this data set (Fig. 2). The second upstream initiation codon was found ten bases upstream (−10 position) but out-of-frame from the cic. This second alternative initation codon (aic) was identified in the matK gene sequence for all 13 orchid taxa examined and is the result of a four base insertion (ATGT) not found in the non-orchid monocots we examined (Fig. 2a, indel 1). Translation using the alternative initiation codon produced a full-length MatK reading frame for eight of the ten orchid taxa previously reported to contain premature stop codons when translated with the cic (Fig. 2b). A single questionable stop codon was found in MatK of Anthosiphon roseans (amino acid position 188) and Caladenia catenata (amino acid position 345) when translated using the aic. It is to be noted that the cic and the aic in Anthosiphon roseans are in the same frame (Fig. 2a).Table 1


Alternative translation initiation codons for the plastid maturase MatK: unraveling the pseudogene misconception in the Orchidaceae.

Barthet MM, Moukarzel K, Smith KN, Patel J, Hilu KW - BMC Evol. Biol. (2015)

Sample alignments showing the consensus initiation codon (cic) and subsequent translation products for MatK in orchids. (a) Nucleotide alignment of various orchid species and the non-orchid monocot species Asparagus aethiopicus, Hordeum vulgare, Saccharum officinarum and Oryza sativa. Asterisks next to species name indicate those noted in GenBank to contain matK as a pseudogene. Black arrow indicates cic used by other monocots and angiosperms for MatK expression. (b) Translated MatK amino acid sequence using the cic. Asterisk in translated amino acid sequence indicates stop codons in the MatK reading frame. Gray shadowing indicates highly conserved sequence among all taxa in the alignment, lighter gray indicates less conserved sequence, white background indicates a lack of conserved sequence. Note: gaps in nucleotide and amino acid alignment differ in relative position due to indel in Palmorchis trilobulata
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

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Fig1: Sample alignments showing the consensus initiation codon (cic) and subsequent translation products for MatK in orchids. (a) Nucleotide alignment of various orchid species and the non-orchid monocot species Asparagus aethiopicus, Hordeum vulgare, Saccharum officinarum and Oryza sativa. Asterisks next to species name indicate those noted in GenBank to contain matK as a pseudogene. Black arrow indicates cic used by other monocots and angiosperms for MatK expression. (b) Translated MatK amino acid sequence using the cic. Asterisk in translated amino acid sequence indicates stop codons in the MatK reading frame. Gray shadowing indicates highly conserved sequence among all taxa in the alignment, lighter gray indicates less conserved sequence, white background indicates a lack of conserved sequence. Note: gaps in nucleotide and amino acid alignment differ in relative position due to indel in Palmorchis trilobulata
Mentions: Two data sets were generated, one for the molecular and the other for the informatics aspects of this study. In the former case, GenBank nucleotide sequences were obtained for 13 orchid and four related monocot species representing cases where matK was considered as either functional or a pseudogene (Table 1). Translation of matK sequence from the initiation codon used for other monocots (referred to hereafter as the consensus initiation codon, cic) produced full length amino acid sequence for MatK protein from the monocot species Asparagus aethiopicus , Hordeum vulgare, Saccharum officinarum, and Oryza sativa , and three orchid species (Fig. 1). Amino acid sequence from the remaining ten orchid species when translated from the cic displayed premature stop codons suggestive of truncated, non-functional protein (Fig. 1b). Two additional initiation codons were identified upstream of the cic. The first one located −6 bp upstream and in-frame with the cic, was found only in six of the thirteen orchid taxa examined in this data set (Fig. 2). The second upstream initiation codon was found ten bases upstream (−10 position) but out-of-frame from the cic. This second alternative initation codon (aic) was identified in the matK gene sequence for all 13 orchid taxa examined and is the result of a four base insertion (ATGT) not found in the non-orchid monocots we examined (Fig. 2a, indel 1). Translation using the alternative initiation codon produced a full-length MatK reading frame for eight of the ten orchid taxa previously reported to contain premature stop codons when translated with the cic (Fig. 2b). A single questionable stop codon was found in MatK of Anthosiphon roseans (amino acid position 188) and Caladenia catenata (amino acid position 345) when translated using the aic. It is to be noted that the cic and the aic in Anthosiphon roseans are in the same frame (Fig. 2a).Table 1

Bottom Line: We confirm that MatK protein is expressed and functions in sample orchids currently described as having a matK pseudogene using immunodetection and reverse-transcription methods.We demonstrate using phylogenetic analysis that this alternative initiation codon emerged de novo within the Orchidaceae, with several reversal events at the basal lineage and deep in orchid history.These findings suggest a novel evolutionary shift for expression of matK in the Orchidaceae and support the function of MatK as a group II intron maturase in the plastid genome of land plants including the orchids.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, Coastal Carolina University, Conway, SC, 29526, USA. mbarthet@coastal.edu.

ABSTRACT

Background: The plastid maturase MatK has been implicated as a possible model for the evolutionary "missing link" between prokaryotic and eukaryotic splicing machinery. This evolutionary implication has sparked investigations concerning the function of this unusual maturase. Intron targets of MatK activity suggest that this is an essential enzyme for plastid function. The matK gene, however, is described as a pseudogene in many photosynthetic orchid species due to presence of premature stop codons in translations, and its high rate of nucleotide and amino acid substitution.

Results: Sequence analysis of the matK gene from orchids identified an out-of-frame alternative AUG initiation codon upstream from the consensus initiation codon used for translation in other angiosperms. We demonstrate translation from the alternative initiation codon generates a conserved MatK reading frame. We confirm that MatK protein is expressed and functions in sample orchids currently described as having a matK pseudogene using immunodetection and reverse-transcription methods. We demonstrate using phylogenetic analysis that this alternative initiation codon emerged de novo within the Orchidaceae, with several reversal events at the basal lineage and deep in orchid history.

Conclusion: These findings suggest a novel evolutionary shift for expression of matK in the Orchidaceae and support the function of MatK as a group II intron maturase in the plastid genome of land plants including the orchids.

Show MeSH