Molecular Evolution Analysis Of Core Loci Of Flavonoid Biosynthesis Pathway

Flower color is a very important character for angiospermsm and is often considered to attract insects for pollination. In nature, about88%of angiosperm flower color pigments are anthocyanins:pelargonidin, cyanidin and delphinidin. Besides anthocyanins, flanonol glycosides and proanthocyanidins-condensed tannins are other two large groups of flavonoids. Various flavonoids participate in determination of organ pigmentation, cross pollination, ultraviolet screening, pests and disease defence, cold acclimation, etc. in plants. Furthermore, various flavonoids also have various health-protective effects in human body, such as anti-oxidation, free radical scavenging, cancer prevention, anti-inflammation, etc. Therefore, phylogenetic study of the flavonoid biosynthetic pathway will help us to understand the molecular mechanisms and modes of this pathway and related traits, promote basic theory studies on pathway evolution, gene function and protein evolution, and provide guide or reference to molecular design and metabolic engineering of this pathway.In this paper, the major research areas are9key enzymes from23plant species with whole genomes sequenced and stored in JGI database (DOE Join Genome Institute). We downloaded their annotated CDS sequences and annotated Protein sequences to build the Blast databases. We extracted CDSs by Blast, detected domains by InterProScan, and used the R for statistical analysis of homologous gene numbers. Then, we analyzed sequence differences between multiple data sets by ClustalO, and reconstructed phylogenetic trees by ProtTest, Phylip and PhyML. After those, we analyzed the selection pressure of branches and loci, performed codon bias analyses using CodonW, and predicted protein physicochemical properties using EMBOSS, SignalP and Wolf PSort analyses. Then, we collected all information and results to perform global analyses, and get results as below:1. Paralog genes encoding upstream flavonoid pathway enzymes have significantly diverged and have shown taxonomy-specific features. When we compared the results of two kinds of alignments (between orthologs and paralogs respectively), we found that the first kind of alignments showed stable results, the second kind of alignment showed unstable results. And this confusion reflected in the upstream gene obviously. But these paralogs can be extracted from the strict Blast. So we consider that the upstream genes of flavonoid pathway showed disvergence among paralogs among differenct taxonomic units. The duplicated copies of the ancestor gene entered different taxonomic ancestors, and the paralogs diverged separately and differently.2. When CHS is with high copy number, in some species, F3’H is also with high copy number. By the method of gene number statistics, we found that CHS in Rosaceae and Leguminosae with high copy number, F3’H may also be with high copy number. In Poaceae, not only CHS and F3’H but also ANR are with high copy number. It is worthy mentioning that this phenomenon appears not only in taxonomic families, but also in each species therein.3. Codon preference is very different between monocots and dicots. Our analysis of codon usage found a great difference between monocots and dicots. Statisticaly, the third position of a codon in monocots biases to GC with a GC/AT ratio of more than10, but dicots bias to AT. Meanwhile, we also found that the third position of a codon in legumes obviously biases to AT, but other dicots still bias a little to GC though not as significantly as monocots.4. For species from different families, flavonoid biosynthetic pathway genes are quite different in GC content.1) All positions in Gramineae are with high GC content, which is in accord with its high GC content at the third position of a codon.2) In legumes the GC content of all loci show little difference, and throughout the metabolic pathways the GC content shows stability.3) In Brassica plants the GC content in the upstream loci of this metabolic pathway is high but low in the downstream loci.4) In the entire metabolic pathway in dicots, Rosaceae has slightly higher GC content, but still lower than Poaceae from monocotyledons. 5. Single-locus molecular evolution can not represent the evolution of the whole metabolic pathway, and single-pathway evolution can not represent the evolution of the whole species. Through the phylogenetic comparison between artificial micro-chromosome method and the species method, we believe that:single-locus molecular evolution can not represent the evolution of a metabolic pathway, even contrary phenomenon can occur, and even for the most critical and core locus. Single-pathway evolution can not represent the evolution of a while species, even contrary phenomenon can occur, and even if it is the most classic and representative pathway.