Cell wall composition and digestibility alterations in Brachypodium distachyon achieved through reduced expression of the UDP-arabinopyranose mutase

Authors: Rancour, David; Hatfield, Ronald; Marita, Jane; ROHR, NICHOLAS - University Of Georgia; SCHMITZ, ROBERT - University Of Georgia

Submitted to: Frontiers in Plant Science
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 5/31/2015
Publication Date: 6/17/2015
Publication URL: https://handle.nal.usda.gov/10113/62079
Citation: Rancour, D.M., Hatfield, R.D., Marita, J.M., Rohr, N.A., Schmitz, R.J. 2015. Cell wall composition and digestibility alterations in Brachypodium distachyon achieved through reduced expression of the UDP-arabinopyranose mutase. Frontiers in Plant Science. 6:446. doi: 10.3389/fpls.2015.00446.

Interpretive Summary: Grass cell walls are complex structures composed primarily of carbohydrates and lignin with small amounts of protein. Understanding how cell walls are constructed has generally relied upon information gained from taking them apart, much like tearing down a building to determine what holds it up. Unfortunately, one has to look at all the pieces (many are changed from how they were originally used) and guess how they were once put together to make a sound structure. In grass cell walls, there is a key component, a phenolic molecule called ferulic acid, that is responsible for cross-linking polysaccharides to each other and to lignin. This cross-linking limits digestibility of the plants. Ferulates are attached to arabinose on arabinoxylans during the construction of cell walls. We proposed that by decreasing the amount of arabinose in the cell wall, the ferulates and cross-linking would also decrease. There is a critical enzyme needed for the formation of arabinose. By suppressing the gene needed for making this enzyme, we were able to decrease the amount of arabinose in the grass Brachypodium. This, in turn, resulted in a decrease in the amount of ferulate in the cell walls and the total amount of cross-linking. With less ferulate, the walls were more digestible, yet the level of decrease was not so great as to hamper the development of the plant. The use of selective gene suppression provides an opportunity to specifically alter one component of the cell wall and to evaluate how this may alter the function of the cell wall. Such approaches may provide the opportunity to modify cell walls without hampering total biomass production to increase digestibility for improved animal performance.

Technical Abstract: Nucleotide-activated sugars are essential substrates for plant cell wall carbohydrate-polymer biosynthetic glycosyltransferase enzymes. The most prevalent sugars in grass cell walls include glucose (Glc), xylose (Xyl), and arabinose (Ara). These sugars are biosynthetically related via the uridine diphospho (UDP)-sugar interconversion pathway, converting UDP-Glc to UDP-Araf. We sought to target and generate UDP-sugar interconversion pathway mutant transgenic Brachypodium distanchyon lines, resulting in cell wall carbohydrate composition changes with improved digestibility characteristics and without grossly compromising plant stature. Both RNAi-mediated gene suppression and constitutive gene expression approaches were performed. Cell walls from 336 T0 transgenic plants with normal appearance were screened for complete carbohydrate composition. RNAi mutants of BdRGP1, a UDP-arabinopyranose mutase, resulted in the largest alteration in cell wall carbohydrate composition with significant decreases in cell wall Ara content with minimal change in plant stature. Five independent RNAi-RGP1 T1 plant lines were used for in-depth analysis of plant cell wall composition and characteristics. Real-time polymerase chain reaction (PCR) analysis indicated that gene expression levels for BdRGP1, BdRGP2, and BdRGP3 were reduced to 15-20% of controls in RNAi-RGP1 plants. Cell wall Ara content was reduced by 23-51% of control levels. No alterations in cell wall Xyl and Glc content were observed. Corresponding decreases in cell wall ferulic acid (FA) and ferulic acid dimers (FA-dimers) were observed. Additionally, cell wall p-coumarates (pCA) were decreased. We demonstrated that the cell wall pCA decrease corresponds to Ara-coupled pCA. Enzyme-mediated digestibility of RNAi-RGP1 Brachypodium cell walls resulted in a near twofold increase of released carbohydrate. Taken together, our results indicated that targeted manipulation of UDP-sugar biosynthesis results in plant biomass with improved digestibility characteristics for forage and bioenergy applications.