Using a reconstructed flavonoid subnetwork to study anthocyanin biosynthesis

Abstract

Flavonoids are ubiquitous secondary plant metabolites that play a variety of roles in the reproduction and protection of plants. Anthocyanin is a major subgroup of flavonoids that assist plants in attracting pollinators and seed dispersers by providing red to blue pigmentation in flowers and fruits. The compounds are water soluble and occur mostly as glucosylated pigments in fruits, leaves and flowers. A self-contained flavonoid subnetwork, consisting of 137 metabolites and 117 reactions, is extracted from the AraCyc database which contains biochemical pathway information of the model plant Arabidopsis thaliana (Arabidopsis). Arabidopsis is used in this study because of the vast metabolic information available for it as well as the fact that it is very similar to most other plants so should make a good representation of flowering plants. Using the stoichiometric matrix to mathematically represent the connections between the reactions, and convex analysis to identify all possible and feasible metabolic routes at a steady state, 199 elementary modes (EMs) or pathways are derived from the subnetwork. Eighty of these lead to the formation of flavonoid compounds. The rest lead to other compounds such as lignin and phenylpropanoid esters, whose association and interplay with flavonoid production is an interesting result to be further investigated. The study uses the subnetwork to investigate the structural functionality of the anthocyanin biosynthetic pathway (ABP). Two anthocyanin compounds, pelargonidin and cyanidin glucosides, are present in Arabidopsis and six EMs lead to their formation. By identifying the enzymes related to the reactions involved in the six EMs, the structural functionality of the enzyme related genes in the ABP are studied. Genes that play important roles in multiple phenotypic traits are identified and their relationship with other EMs in the flavonoid subnetwork investigated. Analysis is also done to identify and study those genes which, when deleted, would result in the non-production of anthocyanin compounds. Emergent properties of the anthocyanin biosynthetic pathway, such as reaction participation and minimal cut sets are used for the investigations. The reaction participation looks at the multiple phenotypic trait of the anthocyanin biosynthetic pathway genes by determining the number of times the enzyme product of each gene appears in the set of 80 elementary modes responsible for the formation of flavonoid compounds. The results show that, in terms of their sequence in the pathway, genes that occur early in the pathway are involved in more EMs, and thus the formation of other flavonoids, than those found later in the ABP, which are more specific to the anthocyanin compounds. A general observation about the flavonoid subnetwork is that it has a remarkably small number of elementary modes compared to other metabolic networks of similar size in other organisms. This suggests a highly constrained network, which could be due to fact that it deals with secondary metabolites. The results are discussed in more detail in the main paper.