Downregulation of Cinnamyl Alcohol Dehydrogenase or Caffeic Acid O-Methyltransferase Leads to Improved Biological Conversion Efficiency in Brachypodium distachyon
Lignin is a significant recalcitrant in the conversion of plant biomass to bioethanol. Cinnamyl alcohol dehydrogenase (CAD) and caffeic acid O-methyltransferase (COMT) catalyze key steps in the pathway of lignin monomer biosynthesis. Brown midrib mutants in Zea mays and Sorghum bicolor with impaired CAD or COMT activity have attracted considerable agronomic interest for their altered lignin composition and improved digestibility. We identified candidate genes encoding CAD and COMT enzymes in the grass model species Brachypodium distachyon and developed transgenic plants overexpressing artificial microRNA designed to silence BdCAD1 or BdCOMT4. Both transgenes caused altered flowering time and stem count and weight. Downregulation of BdCAD1 caused a leaf brown midrib phenotype, the first time this phenotype has been observed in a C3 plant. While acetyl bromide soluble lignin measurements were equivalent in BdCAD1-silenced and wildtype plants, histochemical staining and thioacidolysis indicated a decrease in lignin syringyl units and reduced syringyl/guaiacyl ratio in the transgenic plants. BdCOMT4-downregulated plants exhibited a decrease in total lignin content, a significant reduction of guaiacyl lignin, and a modest reduction of syringyl lignin. Ethanol yield by microbial fermentation was enhanced in both BdCAD1- and BdCOMT4-downregulated plants. These results have elucidated two key genes in the lignin biosynthetic pathway in B. distachyon that, when perturbed, may result in greater biomass yield and bioconversion efficiency
