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A genetic code alteration is a phenotype diversity generator in the human pathogen Candida albicans.

Miranda I, Rocha R, Santos MC, Mateus DD, Moura GR, Carreto L, Santos MA - PLoS ONE (2007)

Bottom Line: We have reconstructed the early stages of the Candida genetic code alteration by engineering tRNAs that partially reverted the identity of serine CUG codons back to their standard leucine meaning.Such genetic code manipulation had profound cellular consequences as it exposed important morphological variation, altered gene expression, re-arranged the karyotype, increased cell-cell adhesion and secretion of hydrolytic enzymes.Our study provides the first experimental evidence for an important role of genetic code alterations as generators of phenotypic diversity of high selective potential and supports the hypothesis that they speed up evolution of new phenotypes.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, Centro de Estudos do Ambiente e do Mar, University of Aveiro, Aveiro, Portugal.

ABSTRACT

Background: The discovery of genetic code alterations and expansions in both prokaryotes and eukaryotes abolished the hypothesis of a frozen and universal genetic code and exposed unanticipated flexibility in codon and amino acid assignments. It is now clear that codon identity alterations involve sense and non-sense codons and can occur in organisms with complex genomes and proteomes. However, the biological functions, the molecular mechanisms of evolution and the diversity of genetic code alterations remain largely unknown. In various species of the genus Candida, the leucine CUG codon is decoded as serine by a unique serine tRNA that contains a leucine 5'-CAG-3'anticodon (tRNA(CAG)(Ser)). We are using this codon identity redefinition as a model system to elucidate the evolution of genetic code alterations.

Methodology/principal findings: We have reconstructed the early stages of the Candida genetic code alteration by engineering tRNAs that partially reverted the identity of serine CUG codons back to their standard leucine meaning. Such genetic code manipulation had profound cellular consequences as it exposed important morphological variation, altered gene expression, re-arranged the karyotype, increased cell-cell adhesion and secretion of hydrolytic enzymes.

Conclusion/significance: Our study provides the first experimental evidence for an important role of genetic code alterations as generators of phenotypic diversity of high selective potential and supports the hypothesis that they speed up evolution of new phenotypes.

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Related in: MedlinePlus

Ambiguous CUG decoding triggered morphogenesis and phenotypic switching.A) Smooth colony morphology of control clones growing on MM-uri-phloxin B (50µg/ml) agar plates. B) Ambiguous pUA15 clones formed long hyphae, even in absence of external inducers, just growing in MM-uri agar plates at 30°C. Similar results were obtained for pUA13 and pUA14 ambiguous clones (data not shown). D). Expression of S. cerevisiae tRNALeu in C. albicans also induced phenotypic switching, which is characterized by transition between different cell-phase forms, namely white-opaque and myceliated-unmyceliated, giving rise to sectored colonies. D) Phenotypic switching was quantified by counting sectored colonies grown in MM-uri, after 7 days of incubation at 30°C. For each plasmid, up to 10 clones were plated and 3000 colonies were screened.
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pone-0000996-g004: Ambiguous CUG decoding triggered morphogenesis and phenotypic switching.A) Smooth colony morphology of control clones growing on MM-uri-phloxin B (50µg/ml) agar plates. B) Ambiguous pUA15 clones formed long hyphae, even in absence of external inducers, just growing in MM-uri agar plates at 30°C. Similar results were obtained for pUA13 and pUA14 ambiguous clones (data not shown). D). Expression of S. cerevisiae tRNALeu in C. albicans also induced phenotypic switching, which is characterized by transition between different cell-phase forms, namely white-opaque and myceliated-unmyceliated, giving rise to sectored colonies. D) Phenotypic switching was quantified by counting sectored colonies grown in MM-uri, after 7 days of incubation at 30°C. For each plasmid, up to 10 clones were plated and 3000 colonies were screened.

Mentions: Interestingly, expression of those S. cerevisiae tRNALeu genes in C. albicans triggered morphogenesis in both solid and liquid media (Figure 4). The pUA15 clones displayed extensive morphological variation (Figure 4B–C) and produced highly heterogenous cell populations containing elongated-ovoid cells, pseudohypha and true hypha (not shown). Some pUA15 clones produced hypha that occupied sectors or entire colonies. Notably, morphological events that gave rise to these phenotypes happened spontaneously without external inducing factors. As expected, control pUA12 and pUA16 clones had homogeneous morphology and formed smooth-white colonies similar to those of untransformed C. albicans CAI4 (Figure 4A). Similar results were obtained with clones pUA13 and pUA14 (data not shown). Apart from morphogenesis, CUG ambiguity also induced phenotypic switching, which is a C. albicans phenotype characterized by reversible induction of opaque or myceliated sectors in white smooth colonies [34]. High frequency of phenotypic switching (63–88%) was obtained for all clones expressing S. cerevisiae leucine tRNAs (pUA13-15), but not for control clones (pUA12 and pUA16) (Figure 4D).


A genetic code alteration is a phenotype diversity generator in the human pathogen Candida albicans.

Miranda I, Rocha R, Santos MC, Mateus DD, Moura GR, Carreto L, Santos MA - PLoS ONE (2007)

Ambiguous CUG decoding triggered morphogenesis and phenotypic switching.A) Smooth colony morphology of control clones growing on MM-uri-phloxin B (50µg/ml) agar plates. B) Ambiguous pUA15 clones formed long hyphae, even in absence of external inducers, just growing in MM-uri agar plates at 30°C. Similar results were obtained for pUA13 and pUA14 ambiguous clones (data not shown). D). Expression of S. cerevisiae tRNALeu in C. albicans also induced phenotypic switching, which is characterized by transition between different cell-phase forms, namely white-opaque and myceliated-unmyceliated, giving rise to sectored colonies. D) Phenotypic switching was quantified by counting sectored colonies grown in MM-uri, after 7 days of incubation at 30°C. For each plasmid, up to 10 clones were plated and 3000 colonies were screened.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC1991585&req=5

pone-0000996-g004: Ambiguous CUG decoding triggered morphogenesis and phenotypic switching.A) Smooth colony morphology of control clones growing on MM-uri-phloxin B (50µg/ml) agar plates. B) Ambiguous pUA15 clones formed long hyphae, even in absence of external inducers, just growing in MM-uri agar plates at 30°C. Similar results were obtained for pUA13 and pUA14 ambiguous clones (data not shown). D). Expression of S. cerevisiae tRNALeu in C. albicans also induced phenotypic switching, which is characterized by transition between different cell-phase forms, namely white-opaque and myceliated-unmyceliated, giving rise to sectored colonies. D) Phenotypic switching was quantified by counting sectored colonies grown in MM-uri, after 7 days of incubation at 30°C. For each plasmid, up to 10 clones were plated and 3000 colonies were screened.
Mentions: Interestingly, expression of those S. cerevisiae tRNALeu genes in C. albicans triggered morphogenesis in both solid and liquid media (Figure 4). The pUA15 clones displayed extensive morphological variation (Figure 4B–C) and produced highly heterogenous cell populations containing elongated-ovoid cells, pseudohypha and true hypha (not shown). Some pUA15 clones produced hypha that occupied sectors or entire colonies. Notably, morphological events that gave rise to these phenotypes happened spontaneously without external inducing factors. As expected, control pUA12 and pUA16 clones had homogeneous morphology and formed smooth-white colonies similar to those of untransformed C. albicans CAI4 (Figure 4A). Similar results were obtained with clones pUA13 and pUA14 (data not shown). Apart from morphogenesis, CUG ambiguity also induced phenotypic switching, which is a C. albicans phenotype characterized by reversible induction of opaque or myceliated sectors in white smooth colonies [34]. High frequency of phenotypic switching (63–88%) was obtained for all clones expressing S. cerevisiae leucine tRNAs (pUA13-15), but not for control clones (pUA12 and pUA16) (Figure 4D).

Bottom Line: We have reconstructed the early stages of the Candida genetic code alteration by engineering tRNAs that partially reverted the identity of serine CUG codons back to their standard leucine meaning.Such genetic code manipulation had profound cellular consequences as it exposed important morphological variation, altered gene expression, re-arranged the karyotype, increased cell-cell adhesion and secretion of hydrolytic enzymes.Our study provides the first experimental evidence for an important role of genetic code alterations as generators of phenotypic diversity of high selective potential and supports the hypothesis that they speed up evolution of new phenotypes.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, Centro de Estudos do Ambiente e do Mar, University of Aveiro, Aveiro, Portugal.

ABSTRACT

Background: The discovery of genetic code alterations and expansions in both prokaryotes and eukaryotes abolished the hypothesis of a frozen and universal genetic code and exposed unanticipated flexibility in codon and amino acid assignments. It is now clear that codon identity alterations involve sense and non-sense codons and can occur in organisms with complex genomes and proteomes. However, the biological functions, the molecular mechanisms of evolution and the diversity of genetic code alterations remain largely unknown. In various species of the genus Candida, the leucine CUG codon is decoded as serine by a unique serine tRNA that contains a leucine 5'-CAG-3'anticodon (tRNA(CAG)(Ser)). We are using this codon identity redefinition as a model system to elucidate the evolution of genetic code alterations.

Methodology/principal findings: We have reconstructed the early stages of the Candida genetic code alteration by engineering tRNAs that partially reverted the identity of serine CUG codons back to their standard leucine meaning. Such genetic code manipulation had profound cellular consequences as it exposed important morphological variation, altered gene expression, re-arranged the karyotype, increased cell-cell adhesion and secretion of hydrolytic enzymes.

Conclusion/significance: Our study provides the first experimental evidence for an important role of genetic code alterations as generators of phenotypic diversity of high selective potential and supports the hypothesis that they speed up evolution of new phenotypes.

Show MeSH
Related in: MedlinePlus