Patterns of variation in the evolution of carotenoid biosynthetic pathway enzymes of higher plants

Cellular metabolic networks depend on the products of many loci for proper functioning. These interrelationships between loci at the phenotypic level raise the question of whether they evolve independently. Previous research has demonstrated that in the anthocyanin pathway, which produces important secondary metabolites in plants, the genes encoding downstream enzymes show an increased rate of change at nonsynonymous sites when compared with upstream loci due to relaxed constraint. To test whether this pattern exists more broadly, we compared a set of 4 genes encoding enzymes of the carotenoid biosynthetic pathway, which produces a set of distinct colored secondary metabolites in plants. Comparisons between copies of phytoene desaturase, zeta-carotene desaturase, lycopene beta-cyclase, and zeaxanthin epoxidase from 6 taxa indicate that the 3 upstream enzymes (phytoene desaturase, zeta-carotene desaturase, and lycopene beta-cyclase) have similar proportions of codons under selective constraint, whereas the most downstream enzyme (zeaxanthin epoxidase) has more codons evolving under relaxed constraint. Overall, nonsynonymous substitution rates appear to be highest for zeaxanthin epoxidase, whereas synonymous substitution rates were highest for the intermediate enzyme lycopene beta-cyclase. Analysis of codon bias shows that only lycopene beta-cyclase may be under slight selection pressure for codon usage. Taken together, these results show that the enzymes of the carotenoid biosynthetic pathway are under strong selective constraint but that the most downstream enzyme is under the least constraint.