Assessing the utility of seed coat-specific promoters to engineer cell wall polysaccharide composition of mucilage

Polysaccharide composition of seed mucilage was successfully modified using three seed coat-specific promoters driving expression of genes encoding cell wall-modifying enzymes. Arabidopsis thaliana seed coat epidermal cells synthesize and secrete large quantities of mucilage, a specialized secondary cell wall composed of cellulose, hemicellulose, and pectin. The composition and structure of mucilage confers its unique properties of expansion, extrusion, and adherence. We are developing seed mucilage as a model to study the biochemical and biological consequences of manipulating cell wall polysaccharides in vivo using cell wall-modifying enzymes. To specifically engineer mucilage composition and avoid altering other cell types, seed coat-specific promoters are required. In this study, we investigated the ability of seed coat-specific promoters from three genes, TESTA-ABUNDANT2 (TBA2), PEROXIDASE36 (PER36), and MUCILAGE-MODIFIED4 (MUM4), to express the cell wall modifying β-galactosidase (BGAL)-encoding gene MUCILAGE-MODIFIED2 (MUM2) and complement the mum2 mutant. The strength of the three promoters relative to one another was found to vary by two to 250 fold, and correlated with their ability to rescue the mum2 mutant phenotype. The strongest of the three promoters, TBA2p, was then used to examine the ability of three MUM2 homologs to complement the mum2 extrusion and cell wall composition phenotypes. The degree of complementation was variable and correlated with the amino acid sequence similarity between the homologous gene products and MUM2. These data demonstrate that all three seed coat-specific promoters can drive expression of genes encoding carbohydrate-active enzymes in a spatial and temporal pattern sufficiently to modify polysaccharide composition in seed mucilage without obvious negative consequences to the rest of the plant.