The molecular evolution and expression of anthocyanin multigene families in Ipomoea purpurea (common morning glory): Pathway evolution

The objective of this study is to compare the rates of evolution, the frequency and timing of gene duplications, and the expression of multigene families in the anthocyanin biosynthetic pathway. Five questions are addressed: (i) Is positive selection driving the maintenance and functional divergence of duplicate genes in the anthocyanin pathway? (ii) Are anthocyanin gene families evolving at similar rates or undergoing duplications at the same time? (iii) Are there any common trends in the pattern or rate of anthocyanin gene evolution? (iv) Are anthocyanin genes coordinately regulated in pigmented and unpigmented plant organs? and (v) What is the molecular basis for the lack of anthocyanin pigmentation in white flowered Ipomoea purpurea (common morning glory) mutants? To address these questions, genomic and cDNA sequences and northern expression data was utilized from the chalcone synthase (CHS), dihydroflavonol reductase (DFR) and flavanone 3-hydroxylase (F3H) gene families. The number of gene family members was determined and gene phylogenies constructed for CHS, DFR and F3H. Coding sequences and, for DFR, potential regulatory motifs were examined to determine whether enzyme function or expression patterns may differ between gene copies. Rates of synonymous and nonsynonymous change were calculated to determine whether rates of evolution vary within and between anthocyanin gene families. The data suggest that positive selection is maintaining duplicate gene copies by driving changes in enzyme function or in patterns of gene expression. The CHS, F3H and DFR genes involved in anthocyanin biosynthesis are more conserved than the copies with unknown functions. Neither the rates of nucleotide substitution nor the episodes of gene recruitment are coupled within the anthocyanin pathway. Northern analysis reveals correlated gene expression in the pathway, although different groups of genes are upregulated in different plant tissues. The expression data also show that white I. purpurea flower mutants result from two independent mutations. Lack of anthocyanin pigment in the a locus mutant is due to transcriptional blockage of the Chs D gene, while the white flowers of the w mutant is due to mutation at a regulatory locus leading to transcriptional blockage of multiple, late-acting anthocyanin genes.