Characterization of Xylogenic irx8 Cell Suspension Cultures that Produce Reduced Lignin and have Alternative Xylan Biosynthesis

Cellulose, xylan, and lignin are the main components of lignocellulosic biomass that is the potential feedstock for biofuel production. Mutation in the Arabidopsis gene GAUT12/IRX8 was previously reported to cause collapsed xylem along with a reduction of a cell wall xylan and a specific pectic fraction. Phylogenetic analysis shows that GAUT12 belongs to the GAUT1-related gene family. In this study, we generated Arabidopsis seed-derived irx8 and wild type suspension cell cultures and induced tracheary element (TE) formation using 2,4-epibrassinolide (BL). Glycome Profiling revealed changes in the wall glycome of wild type and irx8 cells under xylogenic and control conditions. The induced TEs in irx8 cells cultured under xylogenic conditions had slightly bloated shapes and reduced cell wall thickness compared to TEs in wild type cells cultured under the same conditions. Immunolabeling experiments revealed that irx8 TEs lacked a xylan epitope recognized by LM10, a monoclonal antibody that recognizes low-esterified xylan. Both phloroglucinol-HCl staining and lignin composition analyses revealed that lignin production was reduced in irx8 cells grown under xylogenic conditions. Real-time PCR indicated that this correlated with a reduction in lignin biosynthetic gene expression. We investigated gene expression patterns of putative xylan biosynthetic genes via real-time PCR using RNAs from irx8, wild type, and GAUT12-overexpression lines grown on xylogenic and control media. The results led to the identification of genes tightly correlated with TE formation including those that may correlate with GAUT12-associated xylan biosynthesis. These data suggest that GAUT12 affects lignin and xylan biosynthesis during Arabidopsis secondary cell wall formation.