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Title:   Author  item
                                            Sullivan, Michael
  Submitted to: American Society of Plant Biologists Annual Meeting
                                    Publication Type: Abstract Only
                                    Publication Acceptance Date: 2/25/2005
                                    Publication  Date: 7/1/2005
                                    Citation: Sullivan, M.L. 2005. Cloning and characterization of genes involved  in o-diphenol biosynthesis in red clover [abstract]. Plant Biology 2005. p. 212.Interpretive Summary:
                                Technical Abstract: In red clover leaves, the oxidation of abundant o-diphenol caffeic acid derivatives by the enzyme polyphenol oxidase (PPO) prevents post-harvest protein loss, enhancing the quality of this forage when preserved by ensiling. Unfortunately, many important forage crops, such as alfalfa, lack significant levels of both PPO and o-diphenols in their leaves and stems. We are developing strategies to minimize post-harvest protein losses in a variety of forage crops based on the PPO/o-diphenol system. Caffeic acid and its derivatives are intermediates in the synthesis of monolignols, although in some plants like red clover caffeic acid derivatives accumulate to relatively high levels in leaves. Understanding how red clover leaves synthesize high levels of o-diphenols could greatly facilitate engineering similar pathways in crops like alfalfa. Work on Arabidopsis, basil, and other plants is beginning to identify key enzymes involved in the biosynthesis of o-diphenols and related phenylpropanoid compounds. We have isolated cDNA clones from a red clover leaf cDNA library encoding some of these enzymes including phenylalanine ammonia lyase (PAL), cinnamate 4-hydroxylase (C4H), p-coumaroyl CoA ligase (4CL), hydroxycinnamoyl transferase (HCT), and p-coumaric acid 3-hydroxylase (C3H). There is likely overlap of enzymes involved in the biosynthesis of o-diphenol PPO substrates and other important phenols (e.g. monolignols, phytoalexins), so we are currently characterizing the expression patterns of these genes in red clover as they relate to the accumulation of caffeic acid derivatives. The functional roles of the encoded enzymes in o-diphenol biosynthesis will be determined utilizing gene silencing and overexpression experiments in transgenic red clover and alfalfa, respectively.


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