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The Evolutionary Potential of Phenotypic Mutations.

Yanagida H, Gispan A, Kadouri N, Rozen S, Sharon M, Barkai N, Tawfik DS - PLoS Genet. (2015)

Bottom Line: Exploring putative cryptic signals in all 3'-UTRs of yeast genomes, we found that other enzymes related to NADPH production such as pyruvate carboxylase 1 (PYC1) might be prone to peroxisomal localization via cryptic signals.Using laboratory evolution we found that these translational frameshifts are rapidly imprinted via genetic single base deletions occurring within the very same gene location.Thus, genotypes conferring higher phenotypic variability not only meet immediate challenges by unveiling cryptic 3'-UTR sequences, but also boost the potential for future genetic adaptations.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Chemistry, Weizmann Institute of Science, Rehovot, Israel.

ABSTRACT
Errors in protein synthesis, so-called phenotypic mutations, are orders-of-magnitude more frequent than genetic mutations. Here, we provide direct evidence that alternative protein forms and phenotypic variability derived from translational errors paved the path to genetic, evolutionary adaptations via gene duplication. We explored the evolutionary origins of Saccharomyces cerevisiae IDP3 - an NADP-dependent isocitrate dehydrogenase mediating fatty acids ß-oxidation in the peroxisome. Following the yeast whole genome duplication, IDP3 diverged from a cytosolic ancestral gene by acquisition of a C-terminal peroxisomal targeting signal. We discovered that the pre-duplicated cytosolic IDPs are partially localized to the peroxisome owing to +1 translational frameshifts that bypass the stop codon and unveil cryptic peroxisomal targeting signals within the 3'-UTR. Exploring putative cryptic signals in all 3'-UTRs of yeast genomes, we found that other enzymes related to NADPH production such as pyruvate carboxylase 1 (PYC1) might be prone to peroxisomal localization via cryptic signals. Using laboratory evolution we found that these translational frameshifts are rapidly imprinted via genetic single base deletions occurring within the very same gene location. Further, as exemplified here, the sequences that promote translational frameshifts are also more prone to genetic deletions. Thus, genotypes conferring higher phenotypic variability not only meet immediate challenges by unveiling cryptic 3'-UTR sequences, but also boost the potential for future genetic adaptations.

No MeSH data available.


Related in: MedlinePlus

A. gossypii IDP2 gene partially expresses a peroxisomal isoform by phenotypic frameshift mutation.(A) Growth of the ΔIdp3 strain on petroselinate was complemented with the indicated genes in plasmids. S.cer IDP2 shows no growth whereas S.cer IDP3 promotes full growth. Wild-type A.gos IDP2 partially complements, and one T deletion within its 6T repeat increases growth (A.gos IDP2Δt). Inserting a stop codon in the cryptic PTS1 (A.gos IDP2ΔAKL), or silent mutations in the 6T repeat (into TCTTCT; A.gos IDP2+silent), abolished growth. (B) Wild-type A.gos IDP2 and its two control mutants (A.gos IDP2+silent and A.gos IDP2Δt) to express each isoform, were purified and analyzed by ESI-MS in line with liquid chromatography. The initiator Met is removed in the predicted mass. The relative intensities of original and PTS1-carrying isoforms are 72 and 28 ± 3% (n = 3). (C) Fluorescent visualization of cells co-expressing an mCherry tagged with the C-terminal A.gos IDP2 fragment (Fig 1B; the last 11 amino acids and the 3’-UTR ending with AKL*) and a known peroxisomal protein, Pex14, fused to GFP. White arrows show punctate co-staining with the peroxisomal marker. The frame-shifted fragment (A.gos IDP2Δt) having PTS1 within its coding frame was similarly tested. Scale bar, 4 μm.
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pgen.1005445.g002: A. gossypii IDP2 gene partially expresses a peroxisomal isoform by phenotypic frameshift mutation.(A) Growth of the ΔIdp3 strain on petroselinate was complemented with the indicated genes in plasmids. S.cer IDP2 shows no growth whereas S.cer IDP3 promotes full growth. Wild-type A.gos IDP2 partially complements, and one T deletion within its 6T repeat increases growth (A.gos IDP2Δt). Inserting a stop codon in the cryptic PTS1 (A.gos IDP2ΔAKL), or silent mutations in the 6T repeat (into TCTTCT; A.gos IDP2+silent), abolished growth. (B) Wild-type A.gos IDP2 and its two control mutants (A.gos IDP2+silent and A.gos IDP2Δt) to express each isoform, were purified and analyzed by ESI-MS in line with liquid chromatography. The initiator Met is removed in the predicted mass. The relative intensities of original and PTS1-carrying isoforms are 72 and 28 ± 3% (n = 3). (C) Fluorescent visualization of cells co-expressing an mCherry tagged with the C-terminal A.gos IDP2 fragment (Fig 1B; the last 11 amino acids and the 3’-UTR ending with AKL*) and a known peroxisomal protein, Pex14, fused to GFP. White arrows show punctate co-staining with the peroxisomal marker. The frame-shifted fragment (A.gos IDP2Δt) having PTS1 within its coding frame was similarly tested. Scale bar, 4 μm.

Mentions: We thus focused on unraveling the evolutionary origin of PTS1 motif, that is, when and how IDP3’s peroxisomal signal peptide emerged. By the classical Ohno’s model, the key steps towards divergence occur after duplication, and initially as drift, namely not under adaptive selection [10]. Nonetheless, we searched the C-termini and 3’-UTR sequences immediately after the stop codon of the pre-duplication IDP2s, attempting to identify possible starting sequences from which a PTS1 motif may have evolved via few mutations. We discovered intact, putative PTS1 motifs including an adequate stop codon located shortly after the original stop codon, in the 3’-UTRs. Putative PTS1s were found in 4 out of 5 of Saccaromycetaceae species that are phylogenetically closest to S. cerevisiae but not in more distant species including Candida (so-called CTG fungi group; the bottom clade in Fig 1A). However, unlike previously discovered cryptic PTS1 motifs [30,31,38], these cryptic PTS1 motifs relate not to the enzyme’s coding frame but to a +1 frameshift (Fig 1B). Accordingly, when the cryptic PTS1 was revealed in the coding frame by a single nucleotide deletion upstream to the stop codon, A. gossypii IDP2 (A.gos IDP2) enabled growth of the ΔIdp3 strain on petroselinate (Fig 2A).


The Evolutionary Potential of Phenotypic Mutations.

Yanagida H, Gispan A, Kadouri N, Rozen S, Sharon M, Barkai N, Tawfik DS - PLoS Genet. (2015)

A. gossypii IDP2 gene partially expresses a peroxisomal isoform by phenotypic frameshift mutation.(A) Growth of the ΔIdp3 strain on petroselinate was complemented with the indicated genes in plasmids. S.cer IDP2 shows no growth whereas S.cer IDP3 promotes full growth. Wild-type A.gos IDP2 partially complements, and one T deletion within its 6T repeat increases growth (A.gos IDP2Δt). Inserting a stop codon in the cryptic PTS1 (A.gos IDP2ΔAKL), or silent mutations in the 6T repeat (into TCTTCT; A.gos IDP2+silent), abolished growth. (B) Wild-type A.gos IDP2 and its two control mutants (A.gos IDP2+silent and A.gos IDP2Δt) to express each isoform, were purified and analyzed by ESI-MS in line with liquid chromatography. The initiator Met is removed in the predicted mass. The relative intensities of original and PTS1-carrying isoforms are 72 and 28 ± 3% (n = 3). (C) Fluorescent visualization of cells co-expressing an mCherry tagged with the C-terminal A.gos IDP2 fragment (Fig 1B; the last 11 amino acids and the 3’-UTR ending with AKL*) and a known peroxisomal protein, Pex14, fused to GFP. White arrows show punctate co-staining with the peroxisomal marker. The frame-shifted fragment (A.gos IDP2Δt) having PTS1 within its coding frame was similarly tested. Scale bar, 4 μm.
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pgen.1005445.g002: A. gossypii IDP2 gene partially expresses a peroxisomal isoform by phenotypic frameshift mutation.(A) Growth of the ΔIdp3 strain on petroselinate was complemented with the indicated genes in plasmids. S.cer IDP2 shows no growth whereas S.cer IDP3 promotes full growth. Wild-type A.gos IDP2 partially complements, and one T deletion within its 6T repeat increases growth (A.gos IDP2Δt). Inserting a stop codon in the cryptic PTS1 (A.gos IDP2ΔAKL), or silent mutations in the 6T repeat (into TCTTCT; A.gos IDP2+silent), abolished growth. (B) Wild-type A.gos IDP2 and its two control mutants (A.gos IDP2+silent and A.gos IDP2Δt) to express each isoform, were purified and analyzed by ESI-MS in line with liquid chromatography. The initiator Met is removed in the predicted mass. The relative intensities of original and PTS1-carrying isoforms are 72 and 28 ± 3% (n = 3). (C) Fluorescent visualization of cells co-expressing an mCherry tagged with the C-terminal A.gos IDP2 fragment (Fig 1B; the last 11 amino acids and the 3’-UTR ending with AKL*) and a known peroxisomal protein, Pex14, fused to GFP. White arrows show punctate co-staining with the peroxisomal marker. The frame-shifted fragment (A.gos IDP2Δt) having PTS1 within its coding frame was similarly tested. Scale bar, 4 μm.
Mentions: We thus focused on unraveling the evolutionary origin of PTS1 motif, that is, when and how IDP3’s peroxisomal signal peptide emerged. By the classical Ohno’s model, the key steps towards divergence occur after duplication, and initially as drift, namely not under adaptive selection [10]. Nonetheless, we searched the C-termini and 3’-UTR sequences immediately after the stop codon of the pre-duplication IDP2s, attempting to identify possible starting sequences from which a PTS1 motif may have evolved via few mutations. We discovered intact, putative PTS1 motifs including an adequate stop codon located shortly after the original stop codon, in the 3’-UTRs. Putative PTS1s were found in 4 out of 5 of Saccaromycetaceae species that are phylogenetically closest to S. cerevisiae but not in more distant species including Candida (so-called CTG fungi group; the bottom clade in Fig 1A). However, unlike previously discovered cryptic PTS1 motifs [30,31,38], these cryptic PTS1 motifs relate not to the enzyme’s coding frame but to a +1 frameshift (Fig 1B). Accordingly, when the cryptic PTS1 was revealed in the coding frame by a single nucleotide deletion upstream to the stop codon, A. gossypii IDP2 (A.gos IDP2) enabled growth of the ΔIdp3 strain on petroselinate (Fig 2A).

Bottom Line: Exploring putative cryptic signals in all 3'-UTRs of yeast genomes, we found that other enzymes related to NADPH production such as pyruvate carboxylase 1 (PYC1) might be prone to peroxisomal localization via cryptic signals.Using laboratory evolution we found that these translational frameshifts are rapidly imprinted via genetic single base deletions occurring within the very same gene location.Thus, genotypes conferring higher phenotypic variability not only meet immediate challenges by unveiling cryptic 3'-UTR sequences, but also boost the potential for future genetic adaptations.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Chemistry, Weizmann Institute of Science, Rehovot, Israel.

ABSTRACT
Errors in protein synthesis, so-called phenotypic mutations, are orders-of-magnitude more frequent than genetic mutations. Here, we provide direct evidence that alternative protein forms and phenotypic variability derived from translational errors paved the path to genetic, evolutionary adaptations via gene duplication. We explored the evolutionary origins of Saccharomyces cerevisiae IDP3 - an NADP-dependent isocitrate dehydrogenase mediating fatty acids ß-oxidation in the peroxisome. Following the yeast whole genome duplication, IDP3 diverged from a cytosolic ancestral gene by acquisition of a C-terminal peroxisomal targeting signal. We discovered that the pre-duplicated cytosolic IDPs are partially localized to the peroxisome owing to +1 translational frameshifts that bypass the stop codon and unveil cryptic peroxisomal targeting signals within the 3'-UTR. Exploring putative cryptic signals in all 3'-UTRs of yeast genomes, we found that other enzymes related to NADPH production such as pyruvate carboxylase 1 (PYC1) might be prone to peroxisomal localization via cryptic signals. Using laboratory evolution we found that these translational frameshifts are rapidly imprinted via genetic single base deletions occurring within the very same gene location. Further, as exemplified here, the sequences that promote translational frameshifts are also more prone to genetic deletions. Thus, genotypes conferring higher phenotypic variability not only meet immediate challenges by unveiling cryptic 3'-UTR sequences, but also boost the potential for future genetic adaptations.

No MeSH data available.


Related in: MedlinePlus