Down-regulation of four putative arabinoxylan feruloyl transferase genes from family PF02458 reduces ester-linked ferulate content in rice cell walls

Industrial processes to produce ethanol from lignocellulosic materials are available, but improved efficiency is necessary to make them economically viable. One of the limitations for lignocellulosic conversion to ethanol is the inaccessibility of the cellulose and hemicelluloses within the tight cell wall matrix. Ferulates (FA) can cross-link different arabinoxylan molecules in the cell wall of grasses via diferulate and oligoferulate bridges. This complex cross-linking is thought to be a key factor in limiting the biodegradability of grass cell walls and, therefore, the reduction in FA is an attractive target to improve enzyme accessibility to cellulose and hemicelluloses. Unfortunately, our knowledge of the genes responsible for the incorporation of FA to the cell wall is limited. A bioinformatics prediction based on the gene similarities and higher transcript abundance in grasses relative to dicot species suggested that genes from the pfam family PF02458 may act as arabinoxylan feruloyl transferases. We show here that the FA content in the cell walls and the transcript levels of rice genes Os05g08640, Os06g39470, Os01g09010 and Os06g39390, are both higher in the stems than in the leaves. In addition, an RNA interference (RNAi) construct that simultaneously down-regulates transcript levels of these four genes is associated with a significant reduction in FA of the cell walls from the leaves of the transgenic plants relative to the control (19% reduction, P < 0.0001). Therefore, our experimental results in rice support the bioinformatics prediction that members of family PF02458 are involved in the incorporation of FA into the cell wall in grasses

Combining Electrophoretic and Fluorescence Method for Screening Fine Structural Variations Among Lignin Model Polymers Differing in Monomer Composition

Due to the challenges of cell walls (biomass) and its applications in various new technologies, there is a need of rapid and reliable screening of fine variations in lignin structure. The in vitro synthesized lignin model polymers are good experimental system to relate lignin structure/properties with its applications. We used iso-electric focusing electrophoresis (IEF) and fluorescence spectroscopy for screening fine structural variations in lignin model polymers, synthesized from the three lignin monomers, coniferyl alcohol, ferulic acid and p-coumaric acid, mixed in various ratios. The results were related with the thermal behavior of the polymers, revealed by differential scanning calorimetry. Each polymer had characteristic IEF pattern that can be used as its fingerprint. On the basis of the number and intensity of particular bands, it is possible to detect fine differences between polymer patterns, associated with the charge distribution on the polymer fractions. The blue shift of the main fluorescence maximum position of the polymers increased in the same order as temperature of glass transition, i. e. (polymer from coniferyl alcohol)>[polymer from coniferyl alcohol and ferulic acid 9: 1 (w/w)>[(polymer from coniferyl alcohol, ferulic acid and p-coumaric acid 8: 1: 1)>(polymer from coniferyl alcohol and p-coumaric acid 9: 1). The results show that the proposed combination of the fluorescence method and IEF may be used to gain complementary information on fine structural differences among the polymers, and influence of the types and ratios of the monomers building the polymer structure