The wheat Em promoter drives reporter gene expression in embryo and aleurone tissue of transgenic barley and rice

The early methionine (Em) proteins are members of the late embryogenesis abundant (LEA) group of proteins that have been considered to be embryo specific. The ability of a 646-bp wheat Em promoter to control green fluorescent protein (gfp) expression was investigated in transgenic barley and rice. Seeds of transgenic plants expressed gfp in the developing embryo but also in the aleurone layer. The 646-bp Em promoter also directed strong gfp expression in cells comprising the junction between the endosperm transfer cells and cells of the aleurone layer. Em-gfp expression in transgenic barley showed differences in spatial and temporal control when compared with that observed in transgenic rice. Em-gfp expression was also detected in mature aleurone cells of transgenic barley and rice with and without abscisic acid (ABA) treatment. Reverse transcriptase-polymerase chain reaction (RT-PCR) results indicated the presence of Em and Em-homologous transcript in embryo, aleurone and endosperm tissues of wheat and of barley and rice, respectively. These results suggest that Em proteins may be expressed in both the embryo and aleurone during seed development, possibly providing protection against desiccation in these two tissues that survive seed drying. They may also have a similar role in these tissues during germination. The Em promoter from wheat may be useful in the expression of novel genes in cereal grains, as an embryo- and aleurone-specific promoter complementing other available endosperm- and pericarp-specific promoters to collectively increase the expression of transgenes in seeds

Structure-function relationships of beta-D-glucan endo- and exohydrolases from higher plants

(1→3),(1→4)-β-D-Glucans represent an important component of cell walls in the Poaceae family of higher plants. A number of glycoside endo- and exohydrolases is required for the depolymerization of (1→3),(1→4)-β-Dglucans in germinated grain or for the partial hydrolysis of the polysaccharide in elongating vegetative tissues. The enzymes include (1→3),(1→4)-β-D-glucan endohydrolases (EC 3.2.1.73), which are classified as family 17 glycoside hydrolases, (1→4)-β-D-glucan glucohydrolases (family 1) and β-D-glucan exohydrolases (family 3). Kinetic analyses of hydrolytic reactions enable the definition of action patterns, the thermodynamics of substrate binding, and the construction of subsite maps. Mechanism-based inhibitors and substrate analogues have been used to study the spatial orientation of the substrate in the active sites of the enzymes, at the atomic level. The inhibitors and substrate analogues also allow us to define the catalytic mechanisms of the enzymes and to identify catalytic amino acid residues. Three-dimensional structures of (1→3),(1→4)-β-D-glucan endohydrolases, (1→4)- β-D-glucan glucohydrolases and β-D-glucan exohydrolases are available or can be reliably modelled from the crystal structures of related enzymes. Substrate analogues have been diffused into crystals for solving of the threedimensional structures of enzyme-substrate complexes. This information provides valuable insights into potential biological roles of the enzymes in the degradation of the barley (1→3),(1→4)-β-D-glucans during endosperm mobilization and in cell elongation.Maria Hrmova and Geoffrey B. Finche