Adaptive evolution of the chrysanthemyl diphosphate synthase gene involved in irregular monoterpene metabolism

Abstract

Background: Chrysanthemyl diphosphate synthase (CDS) is a key enzyme in biosynthetic pathways producing pyrethrins and irregular monoterpenes. These compounds are confined to plants of the tribe Anthemideae of the Asteraceae, and play an important role in defending the plants against herbivorous insects. It has been proposed that the CDS genes arose from duplication of the farnesyl diphosphate synthase (FDS) gene and have different function from FDSs. However, the duplication time toward the origin of CDS and the evolutionary force behind the functional divergence of the CDS gene are still unknown.

Results: Two duplication events were detected in the evolutionary history of the FDS gene family in the Asteraceae, and the second duplication led to the origin of CDS. CDS occurred after the divergence of the tribe Mutisieae from other tribes of Asteraceae but before the birth of the Anthemideae tribe. After its origin, CDS accumulated four mutations in sites homologous to the substrate-binding and catalysis sites of FDS. Of these, two sites were involved in the binding of the nucleophilic substrate isopentenyl diphosphate in FDS. Maximum likelihood analyses showed that some sites in CDS were under positive selection and were scattered throughout primary sequences, whereas in the three-dimensional structure model they clustered in the large central cavity.

Conclusion: Positive selection associated with gene duplication played a major role in the evolution of CDS.

Figure 1

Phylogenetic relationships of major clades of Asteraceae and phylogeny of the FDS gene family. (A) Phylogenetic relationships of major clades of Asteraceae modified from the Figure 1 of Panero & Funk [41]. Bold branches indicate those most likely for the duplication event in the CDS family. (B) Phylogeny of the FDS gene family reconstructed by the ML method. Numbers next to branches are bootstrap percentages from Maximum Likelihood analysis, and posterior probabilities from Bayesian analysis. The accession numbers of each sequence are given in Additional file 2. The arrows indicate the duplication events. The asterisks indicate sequences that were not used in the codon-based analysis because of their incomplete coding regions. The colors indicate the tribes to which the species belong.

Figure 2

Alignment of deduced amino-acid sequences and three-dimensional models of FDS and CDS. (A) Multiple sequence alignment of FDS and CDS. The amino-acid positions are numbered relative to CDS in P. cinerariifolium (I13995). The consensus sequence highlights the five conserved domains identified in FDS. Sites under positive selection identified by PAML are indicated by asterisks (*sites with p>80%, **sites with p>95%, ***sites with p>99%). Amino-acids involved in the binding and catalysis of substrates [29-31,60] are marked by blue arrows. Red triangles show four sites involved in the binding and catalysis of substrates in FDS and mutated in CDS. Species name abbreviations: Aas, Achillea asiatica; Atr, Artemisia tridentata; Aag, Aster ageratoides; Can, Capsicum annuum; Cas, Centella asiatica; Cin, Cichorium intybus; Cla, Chrysanthemum lavandulifolium; Csc, Cynara scolymus; Glu,Gentiana lutea; Gan, Gerbera anandria; Han, Helianthus annuus; Lvu, Leucanthemum vulgare; Hsa, Homo sapiens (NP_001129294) (The human sequence is included for alignment since it was used as a template in the homology modeling); Mpi, Mentha x piperita; Pci, Pyrethrum cinerariaefolium; Pco Pyrethrum coccineum; Sly, Solanum lycopersicum; Tmo, Taraxacum mongolicum; Tof,Taraxacum officinale. (B) Three-dimensional model of FDS (PDB: 2F8Z). R351 and F239 are involved in IPP-binding in FDS. The red dashed lines represent hydrogen bonds. The yellow sphere indicates water. (C) Distribution of positively-selected sites in the homology model of CDS. CDS has a structural model very similar to FDS: the arrangement of 10 core helices around a large central cavity. The sites indentified as positively-selected (p>80%) were clustered in the large central cavity (shown in pink).