; Experimentally Increased Codon Bias in the Drosophila Adh Gene Leads to an Increase in Larval, But Not Adult, Alcohol Dehydrogenase Activity
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Experimentally Increased Codon Bias in the Drosophila Adh Gene Leads to an Increase in Larval, But Not Adult, Alcohol Dehydrogenase Activity

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Although most amino acids can be encoded by more than one codon, the synonymous codons are not used with equal frequency. This phenomenon is known as codon bias and appears to be a universal feature of genomes. The translational selection hypothesis posits that the use of optimal codons, which match the most abundant species of isoaccepting tRNAs, results in increased translational efficiency and accuracy. Previous work demonstrated that the experimental reduction of codon bias in the Drosophila alcohol dehydrogenase (Adh) gene led to a significant decrease in ADH protein expression. In this study we performed the converse experiment: we replaced seven suboptimal leucine codons that occur naturally in the Drosophila melanogaster Adh gene with the optimal codon. We then compared the in vivo ADH activities imparted by the wild-type and mutant alleles. The introduction of optimal leucine codons led to an increase in ADH activity in third-instar larvae. In adult flies, however, the introduction of optimal codons led to a decrease in ADH activity. There is no evidence that other selectively constrained features of the Adh gene, or its rate of transcription, were altered by the synonymous replacements. These results are consistent with translational selection for codon bias being stronger in the larval stage and suggest that there may be a selective conflict over optimal codon usage between different developmental stages. [PUBLICATION ABSTRACT]

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									Copyright Ó 2010 by the Genetics Society of America
DOI: 10.1534/genetics.109.111294



  Experimentally Increased Codon Bias in the Drosophila Adh Gene Leads
  to an Increase in Larval, But Not Adult, Alcohol Dehydrogenase Activity

               Winfried Hense,* Nathan Anderson,† Stephan Hutter,* Wolfgang Stephan,*
                                 John Parsch* and David B. Carlini†,1
             *Department of Biology II, University of Munich, 82152 Planegg-Martinsried, Germany and †Department of Biology,
                                                American University, Washington, DC 20016
                                                    Manuscript received October 23, 2009
                                                  Accepted for publication December 1, 2009


                                                              ABSTRACT
                Although most amino acids can be encoded by more than one codon, the synonymous codons are not
             used with equal frequency. This phenomenon is known as codon bias and appears to be a universal
             feature of genomes. The translational selection hypothesis posits that the use of optimal codons, which
             match the most abundant species of isoaccepting tRNAs, results in increased translational efficiency and
             accuracy. Previous work demonstrated that the experimental reduction of codon bias in the Drosophila
             alcohol dehydrogenase (Adh) gene led to a significant decrease in ADH protein expression. In this study
             we performed the converse experiment: we replaced seven suboptimal leucine codons that occur
             naturally in the Drosophila melanogaster Adh gene with the optimal codon. We then compared the in vivo
             ADH activities imparted by the wild-type and mutant alleles. The introduction of optimal leucine codons
             led to an increase in ADH activity in third-instar larvae. In adult flies, however, the introduction of optimal
             codons led to a decrease in ADH activity. There is no evidence that other selectively constrained features
             of the Adh gene, or its rate of transcription, were altered by the synonymous replacements. These results
             are consistent with translational selection for codon bias being stronger in the larval stage and suggest that
             there may be a selective conflict over optimal codon usage between different developmental stages.




D    ESPITE the redundancy of the genetic code, synony-
       mous codons are not used with equal frequency—a
phenomenon known as codon bias (Ikemura 1981).
                                                                         genes (Sharp and Li 1986; Duret and Mouchiroud
                                                                         1999) and is significantly higher in the functionally
                                                                         constrained codons of proteins (Akashi 1994). Addi-
Codon bias is apparent in the genomes of a wide array of                 tionally, in Drosophila codon bias is highest in genes
organisms including eubacteria, archaea, and both                        with maximal expression in the larval stage, which is the
unicellular and multicellular eukaryotes; it is essentially              stage requiring the fastest rate of protein synthesis
a universal property of genomes. The two main hypo-                      (Vicario et al. 2008). These observations support the
theses that have been proposed to account for synony-                    hypothesis that codon bias results from natural selec-
mous codon bias are (1) mutational bias (including                       tion for translational accuracy and efficiency (Bulmer
biased gene conversion) and (2) natural selection for                    1991), referred to herein as the translational selection
translational accuracy and/or efficiency (reviewed by                     hypothesis.
Akashi 2001; Duret 2002).                                                   Although there is a substantial body of indirect
   In Drosophila, several lines of evidence suggest that                 evidence for translational selection driving synonymous
codon bias results from natural selection for trans-                     codon usage in Drosophila, direct experimental evi-
lational accuracy and/or efficiency. The lack of a                        dence for the translational selection hypothesis is
significant association between intronic and synony-                      comparatively sparse. Experimental reduction of codon
mous site base composition indicates that mutational                     bias in the leucine codons of the alcohol dehydrogenase
bias cannot account for codon bias (Vicario et al. 2007).                (Adh) gene, the most highly biased codon family in one
Optimal codons, those synonymous codons whose                            of the most highly expressed genes in the Drosophila
usage shows a statistically significant increase in fre-                  melanogaster genome, resulted in a significant reduction
quency with increasing gene expression (Duret and                        in ADH activity (Carlini and Stephan 2003) and
Mouchiroud 1999), tend to match the most abundant                        rendered flies less tolerant to ecologically relevant levels
species of isoaccepting tRNA (Moriyama and Powell                        of environmental ethanol (Carlini 2004). However, to
1997). Codon bias is most extreme in highly expressed                    date no studies have been conducted to examine the
                                                                         functional effects of experimentally increased codon
                                                                         bias in Drosophila. This is a significant consideration,
 1
  Corresponding author: Department of Biology, American University,      because the levels of codon bias observed in the most
Washington, DC 20016. E-mail: carlini@american.edu                       highly expressed genes rarely approach the theoretical

Genetics 184: 547–555 (February 2010)
548                                                    W. Hense et al.

                             
								
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