Exploring Caffeyl-Lignin Biosynthesis in Cleome hassleriana and Polymerization of Caffeyl Alcohol in Arabidopsis thaliana
Description: C-lignin (caffeyl-lignin) is a novel linear lignin polymer found in the seed coats of several non-crop plants, notably Vanilla planifolia (Vanilla), Jatropha Curcas (Jatropha), and Cleome hassleriana (Cleome). C-lignin has several advantages over normal G/S-lignin, found in the majority of lignocellulosic biomass, for valorization in the context of bioprocessing: less cross-linking to cell wall polysaccharides (less recalcitrant biomass), ordered linkages between monomers (homogeneous polymer), and no branching points (linear polymer). These properties make C-lignin an attractive replacement for native lignin in lignocellulosic biomass crops. The seed coats of Cleome hassleriana (Cleome) synthesize G-lignin during early seed maturation, then switch to synthesis of C-lignin during late maturation. This switch to C-lignin in Cleome seed coats is accompanied by loss of caffeoyl-CoA 3-O-methyltransferase (CCoAOMT) and caffeic acid 3-O-methyltransferase (COMT) activities, along with changes in transcript abundance of several lignin related genes. The focus of this research thesis is to understand the biochemical changes leading to C-lignin deposition in Cleome hassleriana seed coats, and to explore the ability of Arabidopsis thaliana seedlings to polymerize caffeyl alcohol to C-lignin. In this thesis, candidate transcripts were implicated in C-lignin biosynthesis by differential gene expression analysis of transcripts in seed coat tissues at 8-18 days after pollination (DAP) and in non-seed coat tissues. Three candidate genes were selected for recombinant expression and their in vitro kinetic properties were measured with potential substrates. Of the three candidates, a cinnamyl alcohol dehydrogenase (ChCAD5) was found to have high transcript levels during C-lignin formation and have a novel preference for converting caffealdehyde to caffeyl alcohol, the precursor of C-lignin. To determine if accumulation of caffeyl alcohol is sufficient for polymerization of C-lignin, Arabidopsis seedlings grown in a xylem induction system were supplied caffeyl alcohol. Polymerization of caffeyl alcohol was not found to occur in this Arabidopsis system, suggesting the ...
Date: December 2019
Creator: Harkleroad, Aaron Djuanell
Partner: UNT Libraries