Analysis of Micronuclear, Macronuclear and cDNA Sequences Encoding the Regulatory Subunit of cAMP‐Dependent Protein Kinase of <i>Euplotes octocarinatus</i>: Evidence for a Ribosomal Frameshift

Tan, Ming; Heckmann, Klaus; Brünen-Nieweler, Claudia

doi:10.1111/j.1550-7408.2001.tb00418.x

Cited by 17 publications

(15 citation statements)

References 35 publications

(63 reference statements)

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“…At these sites, slow recognition of an in-frame sense codon allows the peptidyltRNA bound to the codon immediately upstream to stimulate a ϩ1 shift in frames. This mechanism occurs in the Ty family of yeast retrotransposons (5,15) and in prokaryotic (11) and eukaryotic (1,3,26,27,32,42,43) cellular genes. Members of our laboratory have studied the mechanism of ϩ1 frameshifting in Ty elements for many years and have arrived at a thorough molecular understanding of the process (reviewed by Stahl et al [39]).…”

Section: Resultsmentioning

confidence: 99%

Translational Accuracy during Exponential, Postdiauxic, and Stationary Growth Phases in Saccharomyces cerevisiae

et al. 2004

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When the yeast Saccharomyces cerevisiae shifts from rapid growth on glucose to slow growth on ethanol, it undergoes profound changes in cellular metabolism, including the destruction of most of the translational machinery. We have examined the effect of this metabolic change, termed the diauxic shift, on the frequency of translational errors. Recoding sites are mRNA sequences that increase the frequency of translational errors, providing a convenient reporter of translational accuracy. We found that the diauxic shift causes no overall change in translational accuracy but does cause a strong reduction in the frequency of one type of programmed error: Ty ؉1 frameshifting. Genetic data suggest that this effect may be due to changes in the relative amounts of tRNA participating in translation elongation. We discuss possible implications for expression strategies that use recoding.

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Section: Resultsmentioning

confidence: 99%

Translational Accuracy during Exponential, Postdiauxic, and Stationary Growth Phases in Saccharomyces cerevisiae

et al. 2004

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“…The analysis is based on the alignment of the 50 bp preceding and following the 5Ј-AAA-TA(A/g)-3Ј frameshift motif from the following frameshift sites/genes: the single frameshifts of pEC2 (GenBank accession no. DQ114952), pEC14 (DQ114962), and the three frameshift sites of pEC26 (DQ114969) identified in this study; E. octocarinatus cyclic AMP-dependent protein kinase (AJ238280) (39) The observed 13% frequency of frameshifting somewhat exceeds the value of ϳ7.5% (5 of 67 genes) obtained from a previous survey of genes available in GenBank (22) and provides support for the notion that euplotids possess an extremely high number of genes requiring ϩ1 frameshifts for expression. While there is still considerable uncertainty as to the true percentage of frameshift genes, as a result of the small sample size, the current data provide a 95% confidence interval of 3.7 to 31.7% for the percentage of frameshift genes.…”

Section: Discussionmentioning

confidence: 99%

“…The putative ϩ1-frameshift genes encode the regulatory subunit of cyclic AMP-dependent protein kinase and a nuclear protein kinase of Euplotes octocarinatus (39,40), a La motif protein (p43) in Euplotes aediculatus (1), the Euplotes crassus Tec2 transposon ORF2 protein (11,18), and the reverse transcriptase subunits of telomerase (TERT) in three euplotid species (20,29,42). Since the complete sequences of less than 100 Euplotes genes have been determined, it appears that frameshifting is unusually common in euplotids, with perhaps Ͼ5% of genes requiring a frameshift for expression (reviewed in reference 22).…”

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confidence: 99%

Sequencing of Random Euplotes crassus Macronuclear Genes Supports a High Frequency of +1 Translational Frameshifting

Klobutcher

2005

Eukaryot Cell

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Programmed translational frameshifts have been identified in genes from a broad range of organisms, but typically only a very few genes in a given organism require a frameshift for expression. In contrast, a recent analysis of gene sequences available in GenBank from ciliates in the genus Euplotes indicated that >5% required one or more ؉1 translational frameshifts to produce their predicted protein products. However, this sample of genes was nonrandom, biased, and derived from multiple Euplotes species. To test whether there truly is an abundance of frameshift genes in Euplotes, and to more accurately assess their frequency, we sequenced a random sample of 25 cloned genes/macronuclear DNA molecules from Euplotes crassus. Three new candidate ؉1 frameshift genes were identified in the sample that encode a membrane occupation and recognition nexus (MORN) repeat protein, a C 2 H 2 -type zinc finger protein, and a Ser/Thr protein kinase. Reverse transcription-PCR analyses indicate that all three genes are expressed in vegetatively proliferating cells and that the mRNAs retain the requirement of a frameshift. Although the sample of sequenced genes is relatively small, the results indicate that the frequency of genes requiring frameshifts in E. crassus is between 3.7% and 31.7% (at a 95% confidence interval). The current and past data also indicate that frameshift sites are found predominantly in genes that likely encode nonabundant proteins in the cell.During the translation of an mRNA, a shift in reading frame is usually a catastrophic event, often resulting in a truncated and/or nonfunctional protein. Translational frameshifting is infrequent during the translation of most mRNAs (Ͻ5 ϫ 10 Ϫ5 frameshifts per codon translated [25]), but a number of mRNAs require a frameshift to express a functional protein and appear to have evolved to stimulate frameshifting. Such programmed translational frameshifting (reviewed in references 13 and 32) is relatively rare but is phylogenetically widespread, as examples are known in prokaryotes, eukaryotes, and a number of mobile genetic elements. Sequence elements within the mRNA facilitate programmed frameshifting, and these typically reside at the site of the frameshift, but more distally located sequences can also be involved.A number of genes in ciliated protozoa of the genus Euplotes (class Spirotrichea) have been identified that appear to require a ϩ1 translational frameshift to produce their protein products. The putative ϩ1-frameshift genes encode the regulatory subunit of cyclic AMP-dependent protein kinase and a nuclear protein kinase of Euplotes octocarinatus (39, 40), a La motif protein (p43) in Euplotes aediculatus (1), the Euplotes crassus Tec2 transposon ORF2 protein (11,18), and the reverse transcriptase subunits of telomerase (TERT) in three euplotid species (20,29,42). Since the complete sequences of less than 100 Euplotes genes have been determined, it appears that frameshifting is unusually common in euplotids, with perhaps Ͼ5% of genes requiring a frameshift for ...

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“…Further evidence of functional frameshifting in the genus Euplotes was recently documented for several genes, those encoding the p43 telomerase subunit (1) and two protein kinases, Eopkar (40) and Eondr2 (41). Most recently, Z. Karamysheva et al (submitted for publication) identified two functional ϩ1 frameshift sequences in an E. crassus gene for the telomerase protein TERT.…”

Section: Vol 2 2003mentioning

confidence: 99%

Selection on the Genes of Euplotes crassus Tec1 and Tec2 Transposons: Evolutionary Appearance of a Programmed Frameshift in a Tec2 Gene Encoding a Tyrosine Family Site-Specific Recombinase

Doak

Witherspoon²,

Jahn

et al. 2003

Eukaryot Cell

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The Tec1 and Tec2 transposons of the ciliate Euplotes crassus carry a gene for a tyrosine-type site-specific recombinase. The expression of the Tec2 gene apparently uses a programmed ؉1 frameshift. To test this hypothesis, we first examined whether this gene has evolved under purifying selection in Tec1 and Tec2. Each element carries three genes, and each has evolved under purifying selection for the function of its encoded protein, as evidenced by a dearth of nonsynonymous changes. This distortion of divergence is apparent in codons both 5 and 3 of the frameshift site. Thus, Tec2 transposons have diverged from each other while using a programmed ؉1 frameshift to produce recombinase, the function of which is under purifying selection. What might this function be? Tyrosine-type site-specific recombinases are extremely rare in eukaryotes, and Tec elements are the first known eukaryotic type II transposons to encode a site-specific recombinase. Tec elements also encode a widespread transposase. The Tec recombinase might function in transposition, resolve products of transposition (bacterial replicative transposons use recombinase or resolvase to separate joined replicons), or provide a function that benefits the ciliate host. Transposons in ciliated protozoa are removed from the macronucleus, and it has been proposed that the transposons provide this "excisase" activity.We have studied the evolution of ciliate Tec transposon sequences to compare their evolution in Euplotes crassus to that of TBE1 transposons in Oxytricha ciliates (49) as well as to examine the appearance of a frameshift site in a Tec2 gene (22). We have confirmed that this frameshift has been a functional mechanism, and we demonstrate that this gene and its homolog in Tec1 elements encode a tyrosine-type site-specific recombinase.The Tec and TBE elements encode a DDE transposase (conserved Asp, Asp, Gln residues in the active site) (10) and appear to be type II elements ("cut-and-paste" or "DNAmediated" elements) (5). The evolution of eukaryotic type II transposons has proved difficult to understand: purifying selection for transposase function is not expected to act on the transposase genes of eukaryotic cut-and-paste transposons within a host population and is generally not found (36,37,48,49). However, TBE1 transposons are a notable exception to this expectation. TBE1 transposon genes have evolved under strong selection for the function of their encoded proteins within their host ciliates (49): that is, nonsynonymous-or missense-mutations have been selected against and so removed from the pool of transposons in the host genome, while synonymous mutations have accumulated unabated. The reason for this selection is at present unknown, but it has been proposed that TBE1 transposons are under selection for a specific host function (28,46,49).Ciliated protozoa are unique transposon hosts because they carry two types of nuclei, a micronucleus and a macronucleus. The macronucleus is a terminally differentiated version of the micronucleus and provides a...

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Analysis of Micronuclear, Macronuclear and cDNA Sequences Encoding the Regulatory Subunit of cAMP‐Dependent Protein Kinase of Euplotes octocarinatus: Evidence for a Ribosomal Frameshift

Cited by 17 publications

References 35 publications

Translational Accuracy during Exponential, Postdiauxic, and Stationary Growth Phases in Saccharomyces cerevisiae

Translational Accuracy during Exponential, Postdiauxic, and Stationary Growth Phases in Saccharomyces cerevisiae

Sequencing of Random Euplotes crassus Macronuclear Genes Supports a High Frequency of +1 Translational Frameshifting

Selection on the Genes of Euplotes crassus Tec1 and Tec2 Transposons: Evolutionary Appearance of a Programmed Frameshift in a Tec2 Gene Encoding a Tyrosine Family Site-Specific Recombinase

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